Phenyl-substituted nicotinic ligands, and methods of use thereof

ABSTRACT

Disclosed are compounds and methods of using them to treat a disorder selected from the group consisting of addiction, pain, obesity, schizophrenia, epilepsy, mania and manic depression, anxiety, Alzheimer&#39;s disease, learning deficit, cognition deficit, attention deficit, memory loss, Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD), Parkinson&#39;s disease, Huntington&#39;s disease, Tourette&#39;s syndrome, amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury, dyskinesias, obsessive compulsive disorder, chemical substance abuse, alcoholism, memory deficit, pseudodementia, Ganser&#39;s syndrome, migraine pain, bulimia, premenstrual syndrome or late luteal phase syndrome, tobacco abuse, post-traumatic syndrome, social phobia, chronic fatigue syndrome, premature ejaculation, erectile difficulty, anorexia nervosa, autism, mutism, trichotillomania, hypothermia, and disorders of sleep.

RELATED APPLICATIONS

This application claims the benefit of priority to U.S. ProvisionalPatent Application Ser. No. 61/558,812, filed Nov. 11, 2011; and U.S.Provisional Patent Application Ser. No. 61/647,223, filed May 15, 2012.

GOVERNMENT SUPPORT

This invention was made with government support under grant numbersDA027990 and R03DA025947 awarded by the National Institutes of Health.The government has certain rights in the invention.

BACKGROUND OF THE INVENTION

Neuronal nicotinic acetylcholine receptors (nAChRs) serve a wide rangeof physiological functions and have been implicated in a number ofpathological processes and pharmacological effects of nicotinic drugs.Many of the important in vivo effects of nicotine in the central nervoussystem (CNS) are mediated mainly by the desensitization of nAChRs,specifically α4β2 nAChRs, which are the major nAChR subtype in the CNSand the one most clearly affected (up-regulated) by chronicadministration of nicotine in rats and mice and by smoking in humans.

Sazetidine-A (Saz-A) is a recently reported new nAChR ligand that is aselective α4β2 nAChR desensitizer. U.S. Pat. No. 8,030,300 (incorporatedby reference). Its major in vitro effect is to desensitize α4β2 nAChRswithout affecting either α3β4 or α7 nAChRs. Saz-A shows strong in vivoeffects in animal models, including analgesia, reduction in nicotineself-administration, reduction in alcohol intake, antidepressant-likeactivity, and reversal of attentional impairment.

SUMMARY OF THE INVENTION

Developing novel compounds to target the nicotinic acetylcholinereceptors containing α4 and β2 subunits with desensitization effectscould provide new effective treatments for nicotinic addiction. Thepresent invention is a new class of nAChR ligands that displays highselectivity and picomolar binding affinity for α4β2 nicotinic receptors.Among the novel compounds, 11 high affinity compounds competed for[³H]-epibatidine binding at α4β2 receptors with K_(i) values in therange of 0.031-0.26 nM. These compounds also have other medicinal andpharmacological properties that should make them good candidates as CNSdrugs. The selected lead compound, VMY-2-95, binds with high affinityand potently desensitizes α4β2 receptors. At a dose of 3 mg/kg, VMY-2-95significantly reduced nicotine self-administration in rats. The resultssupport further characterization of VMY-2-95 and its analogs in animalmodels for developing new therapeutics to treat nicotine addiction.

Compounds of the invention may be useful in treating a mammal sufferingfrom aging, addiction, pain, obesity, schizophrenia, epilepsy, mania andmanic depression, anxiety, Alzheimer's disease, learning deficit,cognition deficit, attention deficit, memory loss, Lewy Body Dementia,Attention Deficit Hyperactivity Disorder (ADHD), Parkinson's disease,Huntington's disease, Tourette's syndrome, amyotrophic lateralsclerosis, inflammation, stroke, spinal cord injury, dyskinesias,obsessive compulsive disorder, chemical substance abuse, alcoholism,memory deficit, pseudodementia, Ganser's syndrome, migraine pain,bulimia, premenstrual syndrome or late luteal phase syndrome, tobaccoabuse, post-traumatic syndrome, social phobia, chronic fatigue syndrome,premature ejaculation, erectile difficulty, anorexia nervosa, autism,mutism, trichtillomania, hypothermia, or disorders of sleep; or inimproving cognitive functions and attention.

An aspect of the invention is a compound of formula (I) or apharmaceutically acceptable salt thereof

wherein:R represents, independently for each occurrence, halogen, C1-C6 alkyl,allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, or trifluoro-C1-C4alkyl;R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6 cycloalkyl;R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;m is an integer ranging from 1 to 3;n is an integer selected from 1 and 2; andz is an integer ranging from 0 to 5;

with the proviso that the compound of formula (I) is not

In one embodiment, m is 1.

In one embodiment n is 1.

In one embodiment, z is 0.

In one embodiment, z is 1.

In one embodiment, z is 2.

In one embodiment R represents, independently for each occurrence,halogen, C1-C6 alkyl, C1-C6 alkyloxy, amino, or trifluoro-C1-C4 alkyl.

In one embodiment R represents, independently for each occurrence,halogen, methyl, methoxy, amino, or trifluoromethyl.

In one embodiment R₁ is C1-C6 alkyl.

In one embodiment R₁ is methyl.

In one embodiment R₁ is hydrogen.

In one embodiment R₂, R₃, and R₄ independently represent hydrogen.

In one embodiment the compound is

In one embodiment the compound is selected from the group consisting of:

An aspect of the invention is a compound of formula (II) or apharmaceutically acceptable salt thereof

wherein:R represents, independently for each occurrence, halogen, C1-C6 alkyl,allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, or trifluoro-C1-C4alkyl;R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6 cycloalkyl;R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;

Y is O, S, or N(R^(C));

R^(C) is hydrogen, C1-C6 alkyl, or allyloxycarbonyl;m is an integer ranging from 1 to 3;n is an integer selected from 1 and 2; andz is an integer ranging from 0 to 5.

In one embodiment m is 1.

In one embodiment n is 1.

In one embodiment, z is 0.

In one embodiment, z is 1.

In one embodiment, z is 2.

In one embodiment R represents, independently for each occurrence,halogen, C1-C6 alkyl, C1-C6 alkyloxy, amino, or trifluoro-C1-C4 alkyl.

In one embodiment R represents, independently for each occurrence,halogen, methyl, methoxy, amino, or trifluoromethyl.

In one embodiment R₁ is C1-C6 alkyl.

In one embodiment R₁ is methyl.

In one embodiment R₁ is hydrogen.

In one embodiment R₂, R₃, and R₄ independently represent hydrogen.

An aspect of the invention is a compound of formula (III) or apharmaceutically acceptable salt thereof

wherein:R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6 cycloalkyl; R₂, R₃,and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, or C3-C6cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R₅ represents

Y represents —C≡C— or O;A represents hydrogen, halogen, methyl, C2-C6 alkyl, methoxy, hydroxy,amino, trifluoromethyl, isopropyl, or t-butyl;W represents O, S, or N(R^(D));R^(D) represents hydrogen, C1-C6 alkyl, or allyloxycarbonyl;m is an integer ranging from 1 to 3; andn is an integer selected from 1 and 2.

In one embodiment m is 1.

In one embodiment n is 1.

In one embodiment R₁ is C1-C6 alkyl.

In one embodiment R₁ is methyl.

In one embodiment R₁ is hydrogen.

In one embodiment R₂, R₃, and R₄ independently represent hydrogen.

In one embodiment Y is —C≡C—.

In one embodiment Y is O.

An aspect of the invention is a compound of formula (IV) or apharmaceutically acceptable salt thereof

wherein:R represents, independently for each occurrence, halogen, C1-C6 alkyl,allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, or trifluoro-C1-C4alkyl;R₁ represents

R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R_(h) represents C1-C6 alkyl, C3-C6 cycloalkyl, aryl, or C1-C6 alkylsubstituted with at least one fluorine;m is an integer ranging from 1 to 3; andn is an integer selected from 1 and 2; andz is an integer ranging from 0 to 5.

In one embodiment m is 1.

In one embodiment n is 1.

In one embodiment, z is 0.

In one embodiment, z is 1.

In one embodiment, z is 2.

In one embodiment R represents, independently for each occurrence,halogen, C1-C6 alkyl, C1-C6 alkyloxy, amino, or trifluoro-C1-C4 alkyl.

In one embodiment R represents, independently for each occurrence,halogen, methyl, methoxy, amino, or trifluoromethyl.

In one embodiment R₂, R₃, and R₄ independently represent hydrogen.

An aspect of the invention is a compound of formula (V) or apharmaceutically acceptable salt thereof

wherein:R₁ represents

R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R_(h) represents C1-C6 alkyl, C3-C6 cycloalkyl, aryl, or C1-C6 alkylsubstituted with at least one fluorine;R₅ represents

Y represents —C≡C— or O;A represents hydrogen, halogen, methyl, C2-C6 alkyl, methoxy, hydroxy,amino, trifluoromethyl, isopropyl, or t-butyl;W represents O, S, or N(R^(D));R^(D) represents hydrogen, C1-C6 alkyl, or allyloxycarbonyl;m is an integer ranging from 1 to 3; andn is an integer selected from 1 and 2.

In one embodiment m is 1.

In one embodiment n is 1.

In one embodiment R₂, R₃, and R₄ independently represent hydrogen.

An aspect of the invention is a pharmaceutical composition, comprising acompound of the invention; and a pharmaceutically acceptable carrier.

An aspect of the invention is a method of treating a disorder selectedfrom the group consisting of addiction, pain, obesity, schizophrenia,epilepsy, mania and manic depression, anxiety, Alzheimer's disease,learning deficit, cognition deficit, attention deficit, memory loss,Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD),Parkinson's disease, Huntington's disease, Tourette's syndrome,amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury,dyskinesias, obsessive compulsive disorder, chemical substance abuse,alcoholism, memory deficit, pseudodementia, Ganser's syndrome, migrainepain, bulimia, premenstrual syndrome or late luteal phase syndrome,tobacco abuse, post-traumatic syndrome, social phobia, chronic fatiguesyndrome, premature ejaculation, erectile difficulty, anorexia nervosa,autism, mutism, trichotillomania, hypothermia, and disorders of sleep.The method comprises the step of administering to a subject in needthereof a therapeutically effective amount of a compound of theinvention. In one embodiment the disorder is addiction.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 depicts Scheme 1, Method B for synthesis of certain indicatedcompounds of the invention.

FIG. 2 depicts Scheme 1, Method C for synthesis of certain indicatedcompounds of the invention.

FIG. 3 depicts Scheme 2 for synthesis of certain indicated compounds ofthe invention.

FIG. 4 depicts Scheme 3 for synthesis of compound VMY-2-205 of theinvention.

FIG. 5 is a graph depicting acute VMY-2-95 effects on nicotineself-administration in rats.

FIG. 6 is a graph depicting acute VMY-2-95 mean effects on nicotineself-administration in rats.

FIG. 7 A shows selected natural and synthetic nAChR ligands. B showsdesign strategy for VMY-2-95. C shows the general structures of thepresent series of compounds.

FIG. 8 shows predicted structural models of α4β2 nAChR. Atomic levelinteractions between VMY-2-95 with α4β2 nAChR are pictured. Binding siteresidues are labeled and shown in a stick model rendering.

FIG. 9 Molecular modeling overlay of VMY-2-95 (colored green) with(−)-nicotine (1) varenecline (2) saz-A (3) in the α4β2 nAChR bindingsite. Residues are labeled and shown in a stick model rendering.

FIG. 10 shows acute VMY-2-95 effects on locomotor activity. The graphshows the effect of VMY-2-95 on locomotor activity in the 8-figure mazeat 1 hour (Mean+S.E.M; n=12).

FIG. 11 shows in vitro rat brain tissue protein binding data forpropranolol, varenicline, sazetidine A, and VMY-2-95.

FIG. 12 shows in vitro rat plasma protein binding data for propranolol,varenicline, sazetidine A, and VMY-2-95.

FIG. 13 shows in vitro rat protein binding data comparing brain andplasma.

FIG. 14 shows a comparison of binding affinities of series 1 compoundsfor rat nAChR subtypes with those of Saz-A, varenicline and nicotine.

FIG. 15 continues the data presented in FIG. 14.

FIG. 16 shows binding affinities of series 2 compounds for rat nAChRsubtypes.

FIG. 17 shows the activation and inhibition of nAChR function byVMY-2-95, Saz-A, varenicline and nicotine.

FIG. 18 shows the inhibition of binding by VMY-2-95 at other targets.^(a)The tested default concentration of VMY-2-95 for the primary bindingassays is 10 μM. For each receptor, the value represents mean ofreplicates. The inhibition data was generously provided by the NationalInstitute of Mental Health; Psychoactive Drug Screening Program (NIMHPDSP). Significant inhibition is considered >50%. ^(b)Negativeinhibition represents a stimulation of binding.

FIG. 19 shows a comparison of the binding affinity (Ki) of VMY-2-95 forα4β2 nAChRs with those for other targets.

FIG. 20 shows calculated LE and physicochemical properties of compoundsin series 1.

FIG. 21 shows CHN data for final compounds in series 1-3.

FIG. 22 shows HPLC purity data for compounds 131 and 205.

DETAILED DESCRIPTION OF THE INVENTION

Nicotinic acetylcholine receptors (nAChRs) are pentameric ligand gatedion channels with significant potential as molecular targets for thedesign of drugs to treat a variety of central nervous system disorders.In vertebrates 12 neuronal nAChR subunits have been identified,including nine alpha subunits (α2-α10) and three beta subunits (β2-β4).These subunits may co-assemble as either heteromeric and homomericpentameric receptors, forming a theoretically large array of receptorsubtypes. The predominant nAChRs in the CNS are the heteromeric α4β2*subtype, composed of α4 and β2 (the * indicates that some of thesenAChRs may contain one or more other subunits as well) and the homomericα7 subtype. Certain areas of brain also contain a high density of α3β4*nAChRs, and this subtype appears to be the predominant nAChR in theautonomic nervous system.

Nicotine interacts with α4β2, α4β2α6* and α7 nAChRs in the dopaminergicmesolimbic pathway, which connects the ventral tegmental area of themidbrain and the limbic system via the nucleus accumbens, and theseeffects of nicotine on brain dopaminergic systems are important inreinforcing drug self-administration. The mesolimbic dopamine system isassumed to mediate the pleasurable and rewarding effects of most drugsof abuse, including nicotine. There is convincing evidence fromextensive in vivo studies that mesolimbic α4β2* nAChRs play a pivotalrole in the reinforcement effects of nicotine. Differential activationand desensitization of nAChR subtypes on dopamine and GABA neurons andpossibly glutamate neurons result in stimulated dopamine release in thenucleus accumbens, leading to positive reinforcement of nicotine.

The α4β2 subtype of nAChR serves as an important target for treatingnicotine addiction as well as several other important conditions,including anxiety, depression and cognitive disorders, and thus severalgroups have targeted these nicotinic receptors with a wide array ofcompounds to develop therapies. Many of the new compounds arestructurally related to the natural nAChR ligands, including nicotine,epibatidine and cytisine (FIG. 7).

In 2006, varenicline (FIG. 7) emerged as the newest drug approved by theFDA as a therapeutic aid for smoking cessation. Clinical studiesindicate that varenicline is the most effective drug currently availableto help people quit smoking, but its effects may be temporary for mostpeople. Furthermore, varenicline shows notable adverse side effects,limits the use of this compound as a therapy for smoking cessation.

We previously reported the synthesis and pharmacological properties ofsazetidine-A (Saz-A) (FIG. 7), which potently and selectivelydesensitizes α2 containing nicotinic receptors, especially α4β2 nAChRs,as measured by whole cell currents and ion efflux assays. In animalmodels, Saz-A reduced nicotine self-administration, decreased alcoholintake and improved performance in tests of attention. Saz-A was alsofound to produce behaviors consistent with potential antidepressantand/or antianxiety effects in rats and mice. These promising in vivoresults suggest saz-A is an excellent starting compound for developingadditional potent and subtype-selective drugs that desensitize α4β2nAChRs. Recent in vivo studies showed a low concentration of saz-A inrat brain, suggesting that optimization of saz-A physicochemicalproperties might enhance the in vivo CNS efficacy of this group ofcompounds.

Here, based on our studies of medicinal and pharmacological propertiesof Saz-A, we designed and synthesized a new line of novel nAChR ligands.In pharmacological studies, these compounds showed very promisingproperties that could lead to more effective treatments for nicotineaddiction as well as other conditions that involve nAChRs.

DEFINITIONS

For convenience, certain terms employed in the specification, examples,and appended claims are collected here.

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

The term “agonist” refers to a compound that mimics the action ofnatural transmitter or, when the natural transmitter is not known,causes changes at the receptor complex in the absence of other receptorligands.

The term “antagonist” refers to a compound that binds to a receptorsite, but does not cause any physiological changes unless anotherreceptor ligand is present.

The term “ligand” refers to a compound that binds at the receptor site.

The term “heteroatom” as used herein means an atom of any element otherthan carbon or hydrogen. Preferred heteroatoms are boron, nitrogen,oxygen, phosphorus, sulfur and selenium.

The term “alkyl” refers to the radical of saturated aliphatic groups,including straight-chain alkyl groups, branched-chain alkyl groups,cycloalkyl (alicyclic) groups, alkyl substituted cycloalkyl groups, andcycloalkyl substituted alkyl groups. In preferred embodiments, astraight chain or branched chain alkyl has 30 or fewer carbon atoms inits backbone (e.g., C₁-C₃₀ for straight chain, C₃-C₃₀ for branchedchain), and more preferably 20 or fewer. Likewise, preferred cycloalkylshave from 3-10 carbon atoms in their ring structure, and more preferablyhave 5, 6 or 7 carbons in the ring structure.

Unless the number of carbons is otherwise specified, “lower alkyl” asused herein means an alkyl group, as defined above, but having from oneto ten carbons, more preferably from one to six carbon atoms in itsbackbone structure. Likewise, “lower alkenyl” and “lower alkynyl” havesimilar chain lengths. Preferred alkyl groups are lower alkyls. Inpreferred embodiments, a substituent designated herein as alkyl is alower alkyl.

The term “aralkyl”, as used herein, refers to an alkyl group substitutedwith an aryl group (e.g., an aromatic or heteroaromatic group).

The terms “alkenyl” and “alkynyl” refer to unsaturated aliphatic groupsanalogous in length and possible substitution to the alkyls describedabove, but that contain at least one double or triple bond respectively.

The term “allyl” refers to a substituent with the structural formulaH₂C═CH—CH₂—. The term “allyloxycarbonyl” (or “Alloc”) refers to a commonprotecting group H₂C═CH—CH₂—O—C(O)—.

The term “aryl” as used herein includes 5-, 6- and 7-memberedsingle-ring aromatic groups that may include from zero to fourheteroatoms, for example, benzene, naphthalene, anthracene, pyrene,pyrrole, furan, thiophene, imidazole, oxazole, thiazole, triazole,pyrazole, pyridine, pyrazine, pyridazine and pyrimidine, and the like.Those aryl groups having heteroatoms in the ring structure may also bereferred to as “aryl heterocycles” or “heteroaromatics.” The aromaticring can be substituted at one or more ring positions with suchsubstituents as described above, for example, halogen, azide, alkyl,aralkyl, alkenyl, alkynyl, cycloalkyl, hydroxyl, alkoxyl, amino, nitro,sulfhydryl, imino, amido, phosphonate, phosphinate, carbonyl, carboxyl,silyl, ether, alkylthio, sulfonyl, sulfonamido, ketone, aldehyde, ester,heterocyclyl, aromatic or heteroaromatic moieties, —CF₃, —CN, or thelike. The term “aryl” also includes polycyclic ring systems having twoor more cyclic rings in which two or more carbons are common to twoadjoining rings (the rings are “fused rings”) wherein at least one ofthe rings is aromatic, e.g., the other cyclic rings can be cycloalkyls,cycloalkenyls, cycloalkynyls, aryls and/or heterocyclyls.

The terms ortho, meta and para apply to 1,2-, 1,3- and 1,4-disubstitutedbenzenes, respectively. For example, the names 1,2-dimethylbenzene andortho-dimethylbenzene are synonymous.

The terms “heterocyclyl” or “heterocyclic group” refer to 3- to10-membered ring structures, more preferably 3- to 7-membered rings,whose ring structures include one to four heteroatoms. Heterocycles canalso be polycycles. Heterocyclyl groups include, for example, azetidine,azepine, thiophene, thianthrene, furan, pyran, isobenzofuran, chromene,xanthene, phenoxathiin, pyrrole, imidazole, pyrazole, isothiazole,isoxazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine,isoindole, indole, indazole, purine, quinolizine, isoquinoline,quinoline, phthalazine, naphthyridine, quinoxaline, quinazoline,cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine,pyrimidine, phenanthroline, phenazine, phenarsazine, phenothiazine,furazan, phenoxazine, pyrrolidine, oxolane, thiolane, oxazole,piperidine, piperazine, morpholine, lactones, lactams such asazetidinones and pyrrolidinones, sultams, sultones, and the like. Theheterocyclic ring can be substituted at one or more positions with suchsubstituents as described above, as for example, halogen, alkyl,aralkyl, alkenyl, alkynyl, cycloalkyl, hydroxyl, amino, nitro,sulfhydryl, imino, amido, phosphonate, phosphinate, carbonyl, carboxyl,silyl, ether, alkylthio, sulfonyl, ketone, aldehyde, ester, aheterocyclyl, an aromatic or heteroaromatic moiety, —CF₃, —CN, or thelike.

The terms “polycyclyl” or “polycyclic group” refer to two or more rings(e.g., cycloalkyls, cycloalkenyls, cycloalkynyls, aryls and/orheterocyclyls) in which two or more carbons are common to two adjoiningrings, e.g., the rings are “fused rings”. Rings that are joined throughnon-adjacent atoms are termed “bridged” rings. Each of the rings of thepolycycle can be substituted with such substituents as described above,as for example, halogen, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl,hydroxyl, amino, nitro, sulfhydryl, imino, amido, phosphonate,phosphinate, carbonyl, carboxyl, silyl, ether, alkylthio, sulfonyl,ketone, aldehyde, ester, a heterocyclyl, an aromatic or heteroaromaticmoiety, —CF₃, —CN, or the like.

The term “carbocycle”, as used herein, refers to an aromatic ornon-aromatic ring in which each atom of the ring is carbon.

As used herein, the term “nitro” means —NO₂; the term “halogen”designates —F, —Cl, —Br or —I; the term “sulfhydryl” means —SH; the term“hydroxyl” means —OH; and the term “sulfonyl” means —SO₂—.

The terms “amine” and “amino” are art-recognized and refer to bothunsubstituted and substituted amines, e.g., a moiety that can berepresented by the general formula:

wherein R₉, R₁₀ and R′₁₀ each independently represent a group permittedby the rules of valence.

The term “acylamino” is art-recognized and refers to a moiety that canbe represented by the general formula:

wherein R₉ is as defined above, and R′₁₁ represents a hydrogen, analkyl, an alkenyl or —(CH₂)_(m)—R₈, where m and R₈ are as defined above.

The term “amido” is art recognized as an amino-substituted carbonyl andincludes a moiety that can be represented by the general formula:

wherein R₉, R₁₀ are as defined above. Preferred embodiments of the amidewill not include imides which may be unstable.

The term “alkylthio” refers to an alkyl group, as defined above, havinga sulfur radical attached thereto. In preferred embodiments, the“alkylthio” moiety is represented by one of —S-alkyl, —S-alkenyl,—S-alkynyl, and —S—(CH₂)_(m)—R₈, wherein m and R₈ are defined above.Representative alkylthio groups include methylthio, ethyl thio, and thelike.

The term “carbonyl” is art recognized and includes such moieties as canbe represented by the general formula:

wherein X is a bond or represents an oxygen or a sulfur, and R₁₁represents a hydrogen, an alkyl, an alkenyl, —(CH₂)_(m)—R₈ or apharmaceutically acceptable salt, R′₁₁ represents a hydrogen, an alkyl,an alkenyl or —(CH₂)_(m)—R₈, where m and R₈ are as defined above. WhereX is an oxygen and R₁₁ or R′₁₁ is not hydrogen, the formula representsan “ester”. Where X is an oxygen, and R₁₁ is as defined above, themoiety is referred to herein as a carboxyl group, and particularly whenR₁₁ is a hydrogen, the formula represents a “carboxylic acid”. Where Xis an oxygen, and R′₁₁ is hydrogen, the formula represents a “formate”.In general, where the oxygen atom of the above formula is replaced bysulfur, the formula represents a “thiolcarbonyl” group. Where X is asulfur and R₁₁ or R′₁₁ is not hydrogen, the formula represents a“thiolester.” Where X is a sulfur and R₁₁ is hydrogen, the formularepresents a “thiolcarboxylic acid.” Where X is a sulfur and R₁₁′ ishydrogen, the formula represents a “thiolformate.” On the other hand,where X is a bond, and R₁₁ is not hydrogen, the above formula representsa “ketone” group. Where X is a bond, and R₁₁ is hydrogen, the aboveformula represents an “aldehyde” group.

The terms “alkoxyl” or “alkoxy” as used herein refers to an alkyl group,as defined above, having an oxygen radical attached thereto.Representative alkoxyl groups include methoxy, ethoxy, propyloxy,tert-butoxy and the like. An “ether” is two hydrocarbons covalentlylinked by an oxygen. Accordingly, the substituent of an alkyl thatrenders that alkyl an ether is or resembles an alkoxyl, such as can berepresented by one of —O-alkyl, —O— alkenyl, —O-alkynyl,—O—(CH₂)_(m)—R₈, where m and R₈ are described above.

The abbreviations Me, Et, Ph, Tf, Nf, Ts, Ms represent methyl, ethyl,phenyl, trifluoromethanesulfonyl, nonafluorobutanesulfonyl,p-toluenesulfonyl and methanesulfonyl, respectively. A morecomprehensive list of the abbreviations utilized by organic chemists ofordinary skill in the art appears in the first issue of each volume ofthe Journal of Organic Chemistry; this list is typically presented in atable entitled Standard List of Abbreviations. The abbreviationscontained in said list, and all abbreviations utilized by organicchemists of ordinary skill in the art are hereby incorporated byreference.

Analogous substitutions can be made to alkenyl and alkynyl groups toproduce, for example, aminoalkenyls, aminoalkynyls, amidoalkenyls,amidoalkynyls, iminoalkenyls, iminoalkynyls, thioalkenyls, thioalkynyls,carbonyl-substituted alkenyls or alkynyls.

As used herein, the definition of each expression, e.g., alkyl, m, n,etc., when it occurs more than once in any structure, is intended to beindependent of its definition elsewhere in the same structure.

It will be understood that “substitution” or “substituted with” includesthe implicit proviso that such substitution is in accordance withpermitted valence of the substituted atom and the substituent, and thatthe substitution results in a stable compound, e.g., which does notspontaneously undergo transformation such as by rearrangement,cyclization, elimination, etc.

As used herein, the term “substituted” is contemplated to include allpermissible substituents of organic compounds. In a broad aspect, thepermissible substituents include acyclic and cyclic, branched andunbranched, carbocyclic and heterocyclic, aromatic and nonaromaticsubstituents of organic compounds. Illustrative substituents include,for example, those described herein above. The permissible substituentscan be one or more and the same or different for appropriate organiccompounds. For purposes of this invention, the heteroatoms such asnitrogen may have hydrogen substituents and/or any permissiblesubstituents of organic compounds described herein which satisfy thevalences of the heteroatoms. This invention is not intended to belimited in any manner by the permissible substituents of organiccompounds.

The phrase “protecting group” as used herein means temporarysubstituents which protect a potentially reactive functional group fromundesired chemical transformations. Examples of such protecting groupsinclude esters of carboxylic acids, silyl ethers of alcohols, andacetals and ketals of aldehydes and ketones, respectively. The field ofprotecting group chemistry has been reviewed (Greene, T. W.; Wuts, P. G.M. Protective Groups in Organic Synthesis, 2^(nd) ed.; Wiley: New York,1991).

Certain compounds of the present invention may exist in particulargeometric or stereoisomeric forms. The present invention contemplatesall such compounds, including cis- and trans-isomers, R- andS-enantiomers, diastereomers, (n)-isomers, (L)-isomers, the racemicmixtures thereof, and other mixtures thereof, as falling within thescope of the invention. Additional asymmetric carbon atoms may bepresent in a substituent such as an alkyl group. All such isomers, aswell as mixtures thereof, are intended to be included in this invention.

If, for instance, a particular enantiomer of a compound of the presentinvention is desired, it may be prepared by asymmetric synthesis, it maybe isolated using chiral chromatography methods, or by derivation with achiral auxiliary, where the resulting diastereomeric mixture isseparated and the auxiliary group cleaved to provide the pure desiredenantiomers. Alternatively, where the molecule contains a basicfunctional group, such as amino, or an acidic functional group, such ascarboxyl, diastereomeric salts are formed with an appropriateoptically-active acid or base, followed by resolution of thediastereomers thus formed by fractional crystallization orchromatographic means well known in the art, and subsequent recovery ofthe pure enantiomers.

Contemplated equivalents of the compounds described above includecompounds which otherwise correspond thereto, and which have the samegeneral properties thereof (e.g., functioning as analgesics), whereinone or more simple variations of substituents are made which do notadversely affect the efficacy of the compound in binding to opioidreceptors. In general, the compounds of the present invention may beprepared by the methods illustrated in the general reaction schemes as,for example, described below, or by modifications thereof, using readilyavailable starting materials, reagents and conventional synthesisprocedures. In these reactions, it is also possible to make use ofvariants which are in themselves known, but are not mentioned here.

For purposes of this invention, the chemical elements are identified inaccordance with the Periodic Table of the Elements, CAS version,Handbook of Chemistry and Physics, 67th Ed., 1986-87, inside cover.

Selected Compounds of the Invention

An aspect of the invention is a compound of formula (I) or apharmaceutically acceptable salt thereof

wherein:R represents, independently for each occurrence, halogen, C1-C6 alkyl,allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, or trifluoro-C1-C4alkyl;R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6 cycloalkyl;R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;m is an integer ranging from 1 to 3;n is an integer selected from 1 and 2; andz is an integer ranging from 0 to 5;with the proviso that the compound of formula (I) is not

In one embodiment, m is 1. In one embodiment, m is 2. In one embodiment,m is 3.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, n is 1. In one embodiment in accordancewith any one of the foregoing combinations of limitations, n is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, z is 0. In one embodiment in accordancewith any one of the foregoing combinations of limitations, z is 1. Inone embodiment in accordance with any one of the foregoing combinationsof limitations, z is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R represents, independently for eachoccurrence, halogen, C1-C6 alkyl, C1-C6 alkyloxy, amino, ortrifluoro-C1-C4 alkyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R represents, independently for eachoccurrence, halogen, methyl, methoxy, amino, or trifluoromethyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is C1-C6 alkyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is methyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is hydrogen.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₂, R₃, and R₄ independently representhydrogen.

In one embodiment the compound is

In one embodiment the compound is selected from the group consisting of:

An aspect of the invention is a compound of formula (II) or apharmaceutically acceptable salt thereof

wherein:R represents, independently for each occurrence, halogen, C1-C6 alkyl,allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, or trifluoro-C1-C4alkyl;R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6 cycloalkyl;R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;

Y is O, S, or N(R^(C));

R^(C) is hydrogen, C1-C6 alkyl, or allyloxycarbonyl;m is an integer ranging from 1 to 3;n is an integer selected from 1 and 2; andz is an integer ranging from 0 to 5.

In one embodiment, m is 1. In one embodiment, m is 2. In one embodiment,m is 3.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, n is 1. In one embodiment in accordancewith any one of the foregoing combinations of limitations, n is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, z is 0. In one embodiment in accordancewith any one of the foregoing combinations of limitations, z is 1. Inone embodiment in accordance with any one of the foregoing combinationsof limitations, z is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R represents, independently for eachoccurrence, halogen, C1-C6 alkyl, C1-C6 alkyloxy, amino, ortrifluoro-C1-C4 alkyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R represents, independently for eachoccurrence, halogen, methyl, methoxy, amino, or trifluoromethyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is C1-C6 alkyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is methyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is hydrogen.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₂, R₃, and R₄ independently representhydrogen.

An aspect of the invention is a compound of formula (III) or apharmaceutically acceptable salt thereof

wherein:R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6 cycloalkyl;R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R₅ represents

Y represents —C≡C— or O;A represents hydrogen, halogen, methyl, C2-C6 alkyl, methoxy, hydroxy,amino, trifluoromethyl, isopropyl, or t-butyl;W represents O, S, or N(R^(D));R^(D) represents hydrogen, C1-C6 alkyl, or allyloxycarbonyl;m is an integer ranging from 1 to 3; andn is an integer selected from 1 and 2.

In one embodiment, m is 1. In one embodiment, m is 2. In one embodiment,m is 3.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, n is 1. In one embodiment in accordancewith any one of the foregoing combinations of limitations, n is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is C1-C6 alkyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₁ is methyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₂, R₃, and R₄ independently representhydrogen.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, Y is —C≡C—.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, Y is O.

An aspect of the invention is a compound of formula (IV) or apharmaceutically acceptable salt thereof

wherein:R represents, independently for each occurrence, halogen, C1-C6 alkyl,allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, or trifluoro-C1-C4alkyl;R₁ represents

R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R_(h) represents C1-C6 alkyl, C3-C6 cycloalkyl, aryl, or C1-C6 alkylsubstituted with at least one fluorine;m is an integer ranging from 1 to 3;n is an integer selected from 1 and 2; andz is an integer ranging from 0 to 5.

In one embodiment, m is 1. In one embodiment, m is 2. In one embodiment,m is 3.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, n is 1. In one embodiment in accordancewith any one of the foregoing combinations of limitations, n is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, z is 0. In one embodiment in accordancewith any one of the foregoing combinations of limitations, z is 1. Inone embodiment in accordance with any one of the foregoing combinationsof limitations, z is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R represents, independently for eachoccurrence, halogen, C1-C6 alkyl, C1-C6 alkyloxy, amino, ortrifluoro-C1-C4 alkyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R represents, independently for eachoccurrence, halogen, methyl, methoxy, amino, or trifluoromethyl.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₂, R₃, and R₄ independently representhydrogen.

An aspect of the invention is a compound of formula (V) or apharmaceutically acceptable salt thereof

wherein:R₁ represents

R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R_(h) represents C1-C6 alkyl, C3-C6 cycloalkyl, aryl, or C1-C6 alkylsubstituted with at least one fluorine;R₅ represents

Y represents —C≡C— or O;A represents hydrogen, halogen, methyl, C2-C6 alkyl, methoxy, hydroxy,amino, trifluoromethyl, isopropyl, or t-butyl;W represents O, S, or N(R^(D));R^(D) represents hydrogen, C1-C6 alkyl, or allyloxycarbonyl;m is an integer ranging from 1 to 3; andn is an integer selected from 1 and 2.

In one embodiment, m is 1. In one embodiment, m is 2. In one embodiment,m is 3.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, n is 1. In one embodiment in accordancewith any one of the foregoing combinations of limitations, n is 2.

In one embodiment in accordance with any one of the foregoingcombinations of limitations, R₂, R₃, and R₄ independently representhydrogen.

In various embodiments in accordance with any one of the foregoingcombinations of limitations, R_(h) represents C1-C6 alkyl, C3-C6cycloalkyl, aryl, or C1-C6 alkyl substituted with 1, 2, 3, 4, 5, or 6fluorines.

An aspect of the invention is a pharmaceutical composition, comprising acompound of the invention; and a pharmaceutically acceptable carrier.

An aspect of the invention is a method of treating a disorder selectedfrom the group consisting of addiction, pain, obesity, schizophrenia,epilepsy, mania and manic depression, anxiety, Alzheimer's disease,learning deficit, cognition deficit, attention deficit, memory loss,Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD),Parkinson's disease, Huntington's disease, Tourette's syndrome,amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury,dyskinesias, obsessive compulsive disorder, chemical substance abuse,alcoholism, memory deficit, pseudodementia, Ganser's syndrome, migrainepain, bulimia, premenstrual syndrome or late luteal phase syndrome,tobacco abuse, post-traumatic syndrome, social phobia, chronic fatiguesyndrome, premature ejaculation, erectile difficulty, anorexia nervosa,autism, mutism, trichotillomania, hypothermia, and disorders of sleep.The method comprises the step of administering to a subject in needthereof a therapeutically effective amount of a compound of theinvention. In one embodiment the disorder is addiction. In oneembodiment the disorder is hypothermia.

Dosages

The dosage of any compositions of the present invention will varydepending on the symptoms, age and body weight of the patient, thenature and severity of the disorder to be treated or prevented, theroute of administration, and the form of the subject composition. Any ofthe subject formulations may be administered in a single dose or individed doses. Dosages for the compositions of the present invention maybe readily determined by techniques known to those of skill in the artor as taught herein.

In certain embodiments, the dosage of the subject compounds willgenerally be in the range of about 0.01 ng to about 10 g per kg bodyweight, specifically in the range of about 1 ng to about 0.1 g per kg,and more specifically in the range of about 100 ng to about 10 mg perkg.

An effective dose or amount, and any possible effects on the timing ofadministration of the formulation, may need to be identified for anyparticular composition of the present invention. This may beaccomplished by routine experiment as described herein, using one ormore groups of animals (preferably at least 5 animals per group), or inhuman trials if appropriate. The effectiveness of any subjectcomposition and method of treatment or prevention may be assessed byadministering the composition and assessing the effect of theadministration by measuring one or more applicable indices, andcomparing the post-treatment values of these indices to the values ofthe same indices prior to treatment.

The precise time of administration and amount of any particular subjectcomposition that will yield the most effective treatment in a givenpatient will depend upon the activity, pharmacokinetics, andbioavailability of a subject composition, physiological condition of thepatient (including age, sex, disease type and stage, general physicalcondition, responsiveness to a given dosage and type of medication),route of administration, and the like. The guidelines presented hereinmay be used to optimize the treatment, e.g., determining the optimumtime and/or amount of administration, which will require no more thanroutine experimentation consisting of monitoring the subject andadjusting the dosage and/or timing.

While the subject is being treated, the health of the patient may bemonitored by measuring one or more of the relevant indices atpredetermined times during the treatment period. Treatment, includingcomposition, amounts, times of administration and formulation, may beoptimized according to the results of such monitoring. The patient maybe periodically reevaluated to determine the extent of improvement bymeasuring the same parameters. Adjustments to the amount(s) of subjectcomposition administered and possibly to the time of administration maybe made based on these reevaluations.

Treatment may be initiated with smaller dosages which are less than theoptimum dose of the compound. Thereafter, the dosage may be increased bysmall increments until the optimum therapeutic effect is attained.

The use of the subject compositions may reduce the required dosage forany individual agent contained in the compositions because the onset andduration of effect of the different agents may be complimentary.

Toxicity and therapeutic efficacy of subject compositions may bedetermined by standard pharmaceutical procedures in cell cultures orexperimental animals, e.g., for determining the LD₅₀ and the ED₅₀.

The data obtained from the cell culture assays and animal studies may beused in formulating a range of dosage for use in humans. The dosage ofany subject composition lies preferably within a range of circulatingconcentrations that include the ED₅₀ with little or no toxicity. Thedosage may vary within this range depending upon the dosage formemployed and the route of administration utilized. For compositions ofthe present invention, the therapeutically effective dose may beestimated initially from cell culture assays.

Formulations

The compositions of the present invention may be administered by variousmeans, depending on their intended use, as is well known in the art. Forexample, if compositions of the present invention are to be administeredorally, they may be formulated as tablets, capsules, granules, powdersor syrups. Alternatively, formulations of the present invention may beadministered parenterally as injections (intravenous, intramuscular orsubcutaneous), drop infusion preparations or suppositories. Forapplication by the ophthalmic mucous membrane route, compositions of thepresent invention may be formulated as eyedrops or eye ointments. Theseformulations may be prepared by conventional means, and, if desired, thecompositions may be mixed with any conventional additive, such as anexcipient, a binder, a disintegrating agent, a lubricant, a corrigent, asolubilizing agent, a suspension aid, an emulsifying agent or a coatingagent.

In formulations of the subject invention, wetting agents, emulsifiersand lubricants, such as sodium lauryl sulfate and magnesium stearate, aswell as coloring agents, release agents, coating agents, sweetening,flavoring and perfuming agents, preservatives and antioxidants may bepresent in the formulated agents.

Subject compositions may be suitable for oral, nasal, topical (includingbuccal and sublingual), rectal, vaginal, aerosol and/or parenteraladministration. The formulations may conveniently be presented in unitdosage form and may be prepared by any methods well known in the art ofpharmacy. The amount of composition that may be combined with a carriermaterial to produce a single dose vary depending upon the subject beingtreated, and the particular mode of administration.

Methods of preparing these formulations include the step of bringinginto association compositions of the present invention with the carrierand, optionally, one or more accessory ingredients. In general, theformulations are prepared by uniformly and intimately bringing intoassociation agents with liquid carriers, or finely divided solidcarriers, or both, and then, if necessary, shaping the product.

Formulations suitable for oral administration may be in the form ofcapsules, cachets, pills, tablets, lozenges (using a flavored basis,usually sucrose and acacia or tragacanth), powders, granules, or as asolution or a suspension in an aqueous or non-aqueous liquid, or as anoil-in-water or water-in-oil liquid emulsion, or as an elixir or syrup,or as pastilles (using an inert base, such as gelatin and glycerin, orsucrose and acacia), each containing a predetermined amount of a subjectcomposition thereof as an active ingredient. Compositions of the presentinvention may also be administered as a bolus, electuary, or paste.

In solid dosage forms for oral administration (capsules, tablets, pills,dragees, powders, granules and the like), the subject composition ismixed with one or more pharmaceutically acceptable carriers, such assodium citrate or dicalcium phosphate, and/or any of the following: (1)fillers or extenders, such as starches, lactose, sucrose, glucose,mannitol, and/or silicic acid; (2) binders, such as, for example,carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone,sucrose and/or acacia; (3) humectants, such as glycerol; (4)disintegrating agents, such as agar-agar, calcium carbonate, potato ortapioca starch, alginic acid, certain silicates, and sodium carbonate;(5) solution retarding agents, such as paraffin; (6) absorptionaccelerators, such as quaternary ammonium compounds; (7) wetting agents,such as, for example, acetyl alcohol and glycerol monostearate; (8)absorbents, such as kaolin and bentonite clay; (9) lubricants, such atalc, calcium stearate, magnesium stearate, solid polyethylene glycols,sodium lauryl sulfate, and mixtures thereof and (10) coloring agents. Inthe case of capsules, tablets and pills, the compositions may alsocomprise buffering agents. Solid compositions of a similar type may alsobe employed as fillers in soft and hard-filled gelatin capsules usingsuch excipients as lactose or milk sugars, as well as high molecularweight polyethylene glycols and the like.

A tablet may be made by compression or molding, optionally with one ormore accessory ingredients. Compressed tablets may be prepared usingbinder (for example, gelatin or hydroxypropylmethyl cellulose),lubricant, inert diluent, preservative, disintegrant (for example,sodium starch glycolate or cross-linked sodium carboxymethyl cellulose),surface-active or dispersing agent. Molded tablets may be made bymolding in a suitable machine a mixture of the subject compositionmoistened with an inert liquid diluent. Tablets, and other solid dosageforms, such as dragees, capsules, pills and granules, may optionally bescored or prepared with coatings and shells, such as enteric coatingsand other coatings well known in the pharmaceutical-formulating art.

Liquid dosage forms for oral administration include pharmaceuticallyacceptable emulsions, microemulsions, solutions, suspensions, syrups andelixirs. In addition to the subject composition, the liquid dosage formsmay contain inert diluents commonly used in the art, such as, forexample, water or other solvents, solubilizing agents and emulsifiers,such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethylacetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyleneglycol, oils (in particular, cottonseed, groundnut, corn, germ, olive,castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethyleneglycols and fatty acid esters of sorbitan, and mixtures thereof.

Suspensions, in addition to the subject composition, may containsuspending agents as, for example, ethoxylated isostearyl alcohols,polyoxyethylene sorbitol and sorbitan esters, microcrystallinecellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth,and mixtures thereof.

Formulations for rectal or vaginal administration may be presented as asuppository, which may be prepared by mixing a subject composition withone or more suitable non-irritating excipients or carriers comprising,for example, cocoa butter, polyethylene glycol, a suppository wax or asalicylate, and which is solid at room temperature, but liquid at bodytemperature and, therefore, will melt in the body cavity and release theactive agent. Formulations which are suitable for vaginal administrationalso include pessaries, tampons, creams, gels, pastes, foams or sprayformulations containing such carriers as are known in the art to beappropriate.

Dosage forms for transdermal administration of a subject compositionincludes powders, sprays, ointments, pastes, creams, lotions, gels,solutions, patches and inhalants. The active component may be mixedunder sterile conditions with a pharmaceutically acceptable carrier, andwith any preservatives, buffers, or propellants which may be required.

The ointments, pastes, creams and gels may contain, in addition to asubject composition, excipients, such as animal and vegetable fats,oils, waxes, paraffins, starch, tragacanth, cellulose derivatives,polyethylene glycols, silicones, bentonites, silicic acid, talc and zincoxide, or mixtures thereof.

Powders and sprays may contain, in addition to a subject composition,excipients such as lactose, talc, silicic acid, aluminum hydroxide,calcium silicates and polyamide powder, or mixtures of these substances.Sprays may additionally contain customary propellants, such aschlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, suchas butane and propane.

Compositions of the present invention may alternatively be administeredby aerosol. This is accomplished by preparing an aqueous aerosol,liposomal preparation or solid particles containing the compound. Anon-aqueous (e.g., fluorocarbon propellant) suspension could be used.Sonic nebulizers may be used because they minimize exposing the agent toshear, which may result in degradation of the compounds contained in thesubject compositions.

Ordinarily, an aqueous aerosol is made by formulating an aqueoussolution or suspension of a subject composition together withconventional pharmaceutically acceptable carriers and stabilizers. Thecarriers and stabilizers vary with the requirements of the particularsubject composition, but typically include non-ionic surfactants(Tweens, Pluronics, or polyethylene glycol), innocuous proteins likeserum albumin, sorbitan esters, oleic acid, lecithin, amino acids suchas glycine, buffers, salts, sugars or sugar alcohols. Aerosols generallyare prepared from isotonic solutions.

Pharmaceutical compositions of this invention suitable for parenteraladministration comprise a subject composition in combination with one ormore pharmaceutically-acceptable sterile isotonic aqueous or non-aqueoussolutions, dispersions, suspensions or emulsions, or sterile powderswhich may be reconstituted into sterile injectable solutions ordispersions just prior to use, which may contain antioxidants, buffers,bacteriostats, solutes which render the formulation isotonic with theblood of the intended recipient or suspending or thickening agents.

Examples of suitable aqueous and non-aqueous carriers which may beemployed in the pharmaceutical compositions of the invention includewater, ethanol, polyols (such as glycerol, propylene glycol,polyethylene glycol, and the like), and suitable mixtures thereof,vegetable oils, such as olive oil, and injectable organic esters, suchas ethyl oleate. Proper fluidity may be maintained, for example, by theuse of coating materials, such as lecithin, by the maintenance of therequired particle size in the case of dispersions, and by the use ofsurfactants.

Kits

This invention also provides kits for conveniently and effectivelyimplementing the methods of this invention. Such kits comprise anysubject composition, and a means for facilitating compliance withmethods of this invention. Such kits provide a convenient and effectivemeans for assuring that the subject to be treated takes the appropriateactive in the correct dosage in the correct manner. The compliance meansof such kits includes any means which facilitates administering theactives according to a method of this invention. Such compliance meansinclude instructions, packaging, and dispensing means, and combinationsthereof. Kit components may be packaged for either manual or partiallyor wholly automated practice of the foregoing methods. In otherembodiments involving kits, this invention contemplates a kit includingcompositions of the present invention, and optionally instructions fortheir use.

EXAMPLES

The invention now being generally described, it will be more readilyunderstood by reference to the following examples which are includedmerely for purposes of illustration of certain aspects and embodimentsof the present invention, and are not intended to limit the invention.

Overview & General Procedures Methods, Design Strategy for VMY-2-95 andRational Optimization of New Series of Nicotinic Ligands.

Utilizing a homologous series, we optimized Saz-A by replacing the alkylhydroxyl with a benzene group resulting in the design of VMY-2-95 (FIG.7). We anticipated that VMY-2-95 would have an enhanced CNS profilebased on the physicochemical properties of PSA (34.15), lipophilicity(log P=2.36) and log BB (0.22). From our modeling studies we envisionedthat the benzene ring would provide more rigidity and favorableinteractions with the α4β2 nAChR subtype (FIG. 8). Further modificationson benzene ring in VMY-2-95 provided the compounds in series 1. Thecompounds (95-139) in series 1 were synthesized as shown in FIGS. 1 and2. For routes 1 and 2 (FIGS. 1 and 2), compounds were synthesizedstarting with N-Boc protected Mitsunobu product VMY-2-3. Thisintermediate was subjected to Sonogashira reaction conditions followedby subsequent deprotection of the Boc group to yield the final compoundsin series 1.

To further understand the importance of the azetidine ring,N-substitution and stereochemistry on the binding affinity of ligandsfor nAChRs, modifications of VMY-2-95 were carried out as shown in FIG.7. Analogues VMY-2-161, VMY-2-177 and VMY-2-191 (FIG. 3), weresynthesized by employing the same methodology as shown in FIGS. 1 and 2.The N-methyl azetidine (VMY-2-205, FIG. 4) was prepared by reductivemethylation of the secondary amine with formaldehyde.

General Methods.

Nicotinic acetylcholine receptors (nAChRs) in the neuronal systems arepentameric structures composed of subunits distinct from those found inskeletal muscles. The existence of nine α-subunits (α2-α10) and threeβ-subunits (β2-β4) in the mammalian neuronal systems has been described.

The predominant subtype with high affinity for nicotine is comprised ofthree α-subunits and two β-subunits.

The affinity of compounds of the invention for nAChRs may beinvestigated in three tests for in vitro inhibition of ³H-epibatidinbinding, ³H-α-bungarotoxin binding and ³H-cytisine binding as describedbelow.

Cell Lines and Cell Culture.

The cell line expressing rat α3β4 nAChRs, KXα3β4R2, was establishedpreviously by stably transfecting HEK 293 cells with combinations of ratnAChR α3 and β4 subunit genes. The cell line expressing human α4β2nAChRs, YXα4β2H1, were established recently (Tuan et al., 2012,manuscript in preparation). These cell lines were maintained in minimumessential medium (MEM) supplemented with 10% fetal bovine serum, 100units/mL penicillin G, 100 mg/mL streptomycin and selective antibioticsat 37° C. with 5% CO₂ in a humidified incubator. Tissue culture mediumand antibiotics were obtained from Invitrogen Corporation (Carlsbad,Calif.), unless otherwise stated. Fetal bovine serum and horse serumwere provided by Gemini Bio-Products (Woodland, Calif.).

[³H]-Epibatidine Radioligand Binding Assay.

Stably transfected cell lines, tissue culture conditions, membranepreparation procedures and binding assays were described previously.Briefly, cultured cells at >80% confluence were removed from theirflasks (80 cm²) with a disposable cell scraper and placed in 10 mL of 50mM Tris.HCl buffer (pH 7.4, 4° C.). The cell suspension was centrifugedat 10,000×g for 5 min and the pellet was collected. The cell pellet wasthen homogenized in 10 mL buffer with a polytron homogenizer andcentrifuged at 36,000 g for 10 min at 4° C. The membrane pellet wasresuspended in fresh buffer, and aliquots of the membrane preparationwere used for binding assays. The concentration of [³H]-epibatidine usedwas ˜500 pM for competition binding assays. Nonspecific binding wasassessed in parallel incubations in the presence of 300 μM nicotine.Bound and free ligands were separated by vacuum filtration throughWhatman GF/C filters treated with 0.5% polyethylenimine. Thefilter-retained radioactivity was measured by liquid scintillationcounting. Specific binding was defined as the difference between totalbinding and nonspecific binding. Data from competition binding assayswere analyzed using Prism 5 (GraphPad Software, San Diego, Calif.).

⁸⁶Rb⁺ Efflux Assay.

Functional properties of compounds at nAChRs expressed in thetransfected cells were measured using ⁸⁶Rb⁺ efflux assays as describedpreviously. In brief, cells were plated into 24-well plates coated withpoly-D-lysine. The plated cells were grown at 37° C. for 18 to 24 hourto reach 85-95% confluence. The cells were then incubated in growthmedium (0.5 mL/well) containing ⁸⁶Rb⁺ (2 μCi/mL) for 4 hour at 37° C.The loading mixture was then aspirated, and the cells were washed fourtimes with 1 mL buffer (15 mM HEPES, 140 mM NaCl, 2 mM KCl, 1 mM MgSO₄,1.8 mM CaCl₂, 11 mM Glucose, pH 7.4). One mL of buffer with or withoutcompounds to be tested was then added to each well. After incubation for2 min, the assay buffer was collected for measurements of ⁸⁶Rb⁺ releasedfrom the cells. Cells were then lysed by adding 1 mL of 100 mM NaOH toeach well, and the lysate was collected for determination of the amountof ⁸⁶Rb⁺ that was in the cells at the end of the efflux assay.Radioactivity of assay samples and lysates was measured by liquidscintillation counting. Total loading (cpm) was calculated as the sum ofthe assay sample and the lysate of each well. The amount of ⁸⁶Rb⁺ effluxwas expressed as a percentage of ⁸⁶Rb⁺ loaded. Stimulated ⁸⁶Rb⁺ effluxwas defined as the difference between efflux in the presence and absenceof nicotine. For obtaining EC₅₀ and E_(max) values, stimulation curveswere constructed in which 8 different concentrations of a ligand wereincluded in the assay. For obtaining an IC_(50(10′)) value, inhibitioncurves were constructed in which 8 different concentrations of acompound were applied to cells for 10 min before 100 μM nicotine wasapplied to measure stimulated efflux. EC₅₀, E_(max) and IC_(50(10′))values were determined by nonlinear least-squares regression analyses(GraphPad, San Diego, Calif.).

Binding Assays for Targets Other than nAChRS.

All binding assays for targets other than nAChRs were performed by theNational Institute of Mental Health's Psychoactive Drug ScreeningProgram (PDSP) supported by NIMH grant HHSN-271-2008-00025-C(PI: BryanRoth). For experimental details (K_(i) determinations, receptor bindingprofiles, functional data, MDR1 data, etc. as appropriate) refer to thePDSP web site http://pdsp.med.unc.edu/.

In Vitro Inhibition of ³H-Cytisine Binding.

The predominant subtype with high affinity for nicotine is comprised ofα4 and β2 subunits. nAChRs of the latter type may selectively be labeledby the nicotine ligand ³H-cytisine.

Tissue preparation may be performed at 0-4° C. unless otherwiseindicated. Cerebral cortices from male Wistar rats (150-250 g) may behomogenized for 20 sec in 15 mL Tris, HCl (50 mM, pH 7.4) containing 120mM NaCl, 5 mM KCl, 1 mM MgCl₂ and 2.5 mM CaCl₂ using an Ultra-Turraxhomogenizer. The homogenate may then be centrifuged at 27,000×g for 10min. The supernatant may then be discarded and the pellet resuspended infresh buffer and centrifuged a second time. The final pellet may beresuspended in fresh buffer (35 mL per g of original tissue) and usedfor binding assays.

Aliquots of 500 μL homogenate may be added to 25 μL of test solution and25 μL of ³H-cytisine (1 nM, final concentration), mixed and incubatedfor 90 min at 2° C. Non-specific binding may then be determined using(−)-nicotine (100 μM, final concentration). After incubation the samplesmay be added to 5 mL of ice-cold buffer and poured directly onto WhatmanGF/C glass fiber filters under suction and immediately washed with 2×5mL ice-cold buffer. The amount of radioactivity on the filters may thenbe determined by conventional liquid scintillation counting. Specificbinding is total binding minus non-specific binding.

In Vitro Inhibition of ³H-α-Bungarotoxin Binding Rat Brain.

α-Bungarotoxin is a peptide isolated from the venom of the Elapidaesnake Bungarus multicinctus and has high affinity for neuronal andneuromuscular nicotinic receptors, where it acts as a potent antagonist.³H-α-Bungarotoxin binds to a single site in rat brain with a uniquedistribution pattern in rat brain.

³H-α-Bungarotoxin labels nAChR are formed by the α7 subunit isoformfound in the brain and the isoform in the neuromuscular junction.Functionally, the α7 homo-oligomer expressed in oocytes has a calciumpermeability greater than neuromuscular receptors and, in some instancesgreater than NMDA channels.

Tissue Preparation.

Preparations may be performed at 0-4° C. unless otherwise indicated.Cerebral cortices from male Wistar rats (150-250 g) may be homogenizedfor 10 sec in 15 mL 20 mM Hepes buffer containing 118 mM NaCl, 4.8 mMKCl, 1.2 mM MgSO₄ and 2.5 mM CaCl₂ (pH 7.5) using an Ultra-Turraxhomogenizer. The tissue suspension may then be centrifuged at 27,000×gfor 10 min. The supernatant is discarded and the pellet is washed twiceby centrifugation at 27,000×g for 10 min in 20 mL fresh buffer, and thefinal pellet may be resuspended in fresh buffer containing 0.01% BSA (35mL per g of original tissue) and used for binding assays.

Aliquots of 500 μL homogenate may be added to 25 μL of test solution and25 μL of ³H-α-bungarotoxin (2 nM, final concentration), mixed andincubated for 2 h at 24° C. Non-specific binding may then be determinedusing (−)-nicotine (1 mM, final concentration). After incubation thesamples may be added to 5 mL of ice-cold Hepes buffer containing 0.05%PEI and poured directly onto Whatman GF/C glass fiber filters (presoakedin 0.1% PEI for at least 6 h) under suction and immediately washed with2×5 mL ice-cold buffer. The amount of radioactivity on the filters maythen be determined by conventional liquid scintillation counting.Specific binding is total binding minus non-specific binding.

In Vitro Inhibition of ³H-Epibatidin Binding.

Epibatidin is an alkaloid that was first isolated from the skin of theEcuadoran frog Epipedobates tricolor and was found to have very highaffinity for neuronal nicotinic receptors, where it acts as a potentagonist. It is believed that ³H-epibatidin binds to sites in rat brain,both of which have pharmacological profiles consistent with neuronalnicotinic receptors and a similar brain regional distribution (Houglinget al., Mol. Pharmacol. 48, 280-287 (1995)).

The high affinity binding sites in rat brain for ³H-epibatidin aremainly α4β2 nAChR subtype.

Tissue Preparation.

Preparations may be performed at 0-4° C. unless otherwise indicated. Theforebrain (cerebellum) from a male Wistar rat (150-250 g) may behomogenized for 10-20 sec in 20 mL Tris, HCl (50 mM, pH 7.4) using anUltra-Turrax homogenizer. The tissue suspension may then be centrifugedat 27,000×g for 10 min. The supernatant is then discarded and the pelletmay then be washed three times by centrifugation at 27,000×g for 10 minin 20 mL fresh buffer, and the final pellet may be resuspended in freshbuffer (400 mL per g of original tissue) and used for binding assays.

Aliquots of 2.0 mL homogenate may be added to 0.100 mL of test solutionand 0.100 mL of ³H-epibatidin (0.3 nM, final concentration), mixed andincubated for 2 h at room temperature. Non-specific binding may then bedetermined using (−)-nicotine (30 μM, final concentration). Afterincubation the samples may then be poured directly onto Whatman GF/Cglass fiber filters (presoaked in 0.1% PET for at least 20 min) undersuction and immediately washed with 2×5 mL ice-cold buffer. The amountof radioactivity on the filters may be determined by conventional liquidscintillation counting. Specific binding is total binding minusnon-specific binding.

General Procedures for Animal Behavioral Studies.

Separate sets of young adult female Sprague-Dawley rats were used forthe nicotine self-administration study (N=15) and for the locomotoractivity study (N=12). The studies were conducted in accordance with theregulations outlined by the Duke University Animal Care and UseCommittee. The rats housed in approved standard laboratory conditions ina Duke University vivarium facility near the testing room to minimizestress induced by transporting the rats. The rats were kept on a 12:12reverse day/night cycle, so that they were in their active phase duringbehavioral testing. The rats in the drug iv self-administration studieswere singly housed to prevent them from damaging each other's catheters.The rats in the locomotor activity studies were housed in groups of 2-3.All rats were allowed access to water at all times; the rats in thenicotine-self-administration study were fed daily approximately 30minutes after completing the sessions while those in the locomotoractivity study had continuous access to food.

VMY-2-95 Administration.

VMY-2-95 was injected s.c. 10 minutes before testing in a volume of 1mL/kg of saline. The doses (0, 0.3, 1 and 3 mg/kg) were given in acounterbalanced order with at least two days between successiveinjections. For the nicotine self-administration study the VMY-2-95 theacute dose-effect study was tested twice while for the locomotoractivity study the dose-effect function was tested once.

Nicotine Self-Administration.

Before beginning nicotine self-administration, the rats were trained forthree sessions on lever pressing for food reinforcement. Then they werefitted with i.v. catheters and they received nicotine infusions (0.03mg/kg/infusion) on an FR1 schedule for ten sessions. The rats weretrained to self-administer nicotine (0.03 mg/kg/infusion, IV) viaoperant lever response (FR1) with a visual secondary reinforcer. Twolevers were available to be pressed and only one caused the delivery ofnicotine on an FR1 schedule. Pressing the lever on the active sideresulted in the activation of the feedback tone for 0.5 second and theimmediate delivery of one 50-μl infusion of nicotine in less than 1second. Each infusion was immediately followed by a one-minute period inwhich the cue lights went out, the house light came on and responseswere recorded but not reinforced.

Locomotor Activity.

Another set of rats (N=12) was tested for acute VMY-2-95 effects onlocomotor activity in a figure-8 maze over the course of a 1-hoursession (see FIG. 10). The mazes had continuous enclosed alleys 10×10 cmin the shape of a figure-8 (Crofton et al., 1991). The dimensions of theapparatus were 70 cm long and 42 cm wide, with a 21×16 cm central arena,a 20-cm high ceiling and two blind alleys extending 20 cm from eitherside. Eight infrared photobeams, which crossed the alleys, indexedlocomotor activity. One photobeam was located on each of the two blindalleys and three were located on each of two loops of the figure-8.Numbers of photobeam breaks were recorded for five minutes blocks overthe one-hour session. The repeated measures were VMY-2-95 dose and therepeated administration of each dose. Significant interactions werefollowed up by tests of the simple main effects. Alpha of p<0.05(two-tailed) was used as the threshold for significance.

Physicochemical Properties and Ligand Efficiency.

Molecular properties of compounds in series 1 were calculated accordingthe available software tools. Ligand binding efficiency was calculatedaccording to the Hopkins equation: LE=1.372*(−log Ki (Moles))/N.Molecular weight and c Log P were calculated from chembiodraw ultra11.0. Polar Surface Area (PSA) was calculated from www.chemicalize.org.Log BB was calculated from the following equation: LogBB=−0.0148PSA+0.132c Log P+0.139.

Chemical Synthesis of Compounds.

All reagents and solvents were commercially available and used withoutfurther purification. Chromatography was performed for purification offinal compounds using a Biotage SP-1 system with silica gel cartridges.NMR spectra were recorded on a Varian 400 MR spectrometer at 400 MHz forH-1 and 100 MHz for C-13. Chemical shifts (6) are given in ppm downfieldfrom tetramethylsilane, and coupling constants (J-values) are reportedin hertz (Hz). Mass spectroscopy was performed using Waters Q-TOFpremier mass spectrometer. The purity of final compounds was evaluatedby CHN analysis.

The purity of final compounds was evaluated by C, H, N analysis(Atlantic microlabs) (FIG. 21) and HPLC methods for compounds 131 and205 FIG. 22). Detailed synthetic and characterization data (series 2 &3) and experimental procedure for computational studies are presentedbelow.

General Procedure for Mitsunobu Reaction (Method A).

To a mixture of 5-Bromo-3-pyridinol (1.2 equiv) and Ph₃P (1.6 equiv) inanhydrous THF taken in a flame-dried flask under N₂, N-Boc protectedalcohol (1 equiv) was added and the mixture was cooled to −10° C.Diethyl azodicarboxylate (40% w/v) in toluene (1.6 equiv) was addeddropwise to the mixture and was warmed gradually to the roomtemperature. After 48 h, the reaction mixture was quenched with 1 mL ofwater and the solvent was removed under reduced pressure. The resultingyellow oil was purified by column chromatography on silica gel to yield55-60% as a white solid.

General Procedure for Sonogashira Coupling Reaction. One-Pot Synthesis(Method B):

The Mitsunobu adduct (1 equiv), Pd(PPh₃)₂Cl₂ (0.04 eq, 4 mol %), CuI(0.08 eq, 8 mol %), and PPh₃ (0.08 eq, 8 mol %) was placed in anoven-dried round bottom flask with nitrogen. After addition of i-Pr₂NH(1 mL) and toluene (3-5 mL), the mixture was stirred at room temperaturefor 5 min and (trimethylsilyl) acetylene (2.7 equiv) was added andstirred at r.t for 10 min. The whole reaction mixture was stirred at 80°C. for 18 h, a solution of KOH in methanol and water (4:1) was added inone portion and the mixture was stirred for additional 3 h at 25° C.Then the second substituted aryl iodide was added and stirred continuedfor 16 h at 25° C. or 80° C. in case of substituted aryl bromide. Thereaction mixture was quenched with saturated NH₄Cl solution andextracted with CH₂Cl₂. The combined organic layers were washed with 2 NHCl, water and saturated NaCl solution. The organic phase was separatedand dried over anhydrous sodium sulfate, filtered, and concentratedunder reduced pressure to give crude product. The crude product waspurified by column chromatography. See FIG. 1.

Sequential Desilylation and Sonogashira Coupling of TMS-ProtectedVMY-2-87. (Method C).

An oven-dried and nitrogen-filled round bottom flask was charged with(S)-tert-butyl-2-((5-((trimethylsilyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-87, 1 equiv), KOH (2 equiv) methanol in water (20:1). The wholereaction mixture was stirred 25° C. for 3 h. It was then added to asecond flask which contained a performed mixture of Pd(PPh₃)₂Cl₂ (0.02eq, 2 mol %), CuI (0.02 eq, 2 mol %), PPh₃ (0.04 eq, 4 mol %) i-Pr₂NH (1mL), toluene (3-5 mL) and substituted fluoro iodobenzene that hadalready stirred at 25° C. for 30 min. The complete mixture was thenstirred at 25° C. for 16 h. The reaction mixture was quenched withsaturated NH₄Cl solution and extracted with CH₂Cl₂. The combined organiclayers were washed with 2 N HCl, water and saturated NaCl solution. Theorganic phase was separated and dried over anhydrous sodium sulfate,filtered, and concentrated under reduced pressure to give crude product.The crude product was purified by column chromatography. See FIG. 2.

General Procedure for Deprotection of the N-Boc Precursors (Method D).

To a stirred solution of N-Boc protected compound in dichloromethane wasadded trifluoroacetic acid (5-10 equiv) dropwise at 0° C. under anitrogen atmosphere. The reaction mixture was stirred at roomtemperature for 4-6 h (TLC showed complete deprotection of Boc after 5h). The solvent and excess of TFA were removed under reduced pressure.The resulting residue was flushed with nitrogen to remove the traces ofTFA, and was taken up in 2-3 mL methanol followed by dropwise additionof 2 M NaOH solution in methanol and water (9:1) at 0° C. until the pHreached 9-10. After the reaction mixture stirred for 18 h, the reactionmixture was taken in dichloromethane and evaporated. The resultingresidue was purified by column chromatography.

Example 1

Synthesis of(S)-3-((1-methylazetidin-2-yl)methoxy)-5-(phenylethynyl)pyridine(VMY-2-205)

(S)-3-(azetidin-2-ylmethoxy)-5-(phenylethynyl)pyridine (VMY-2-95, 0.12mmol) was taken up in 2 mL of ethanol. Formalin (37%, 0.3 mL) was added,and the acidity was adjusted to pH 5 with the addition of acetic acidand sodium acetate. The reaction mixture was stirred for 15 min. Sodiumcyanoborohydride (0.38 mmol) was added. The whole reaction mixture wasallowed to stir for 18 h at room temperature. The solvent was evaporatedand the crude product was purified by column chromatography to yieldpure VMY-2-205 as a liquid (20 mg, 48%). ¹H NMR (399 MHz, CDCl3) δ 8.30(d, J=1.4, 1H), 8.20 (d, J=2.8, 1H), 7.52-7.43 (m, 2H), 7.32-7.28 (m,3H), 7.25 (dd, J=2.7, 1.7, 1H), 4.06-3.95 (m, 2H), 3.51-3.35 (m, 2H),2.86 (dd, J=15.9, 8.6, 1H), 2.37 (s, 3H), 2.05 (td, J=8.7, 6.0, 2H). ¹³CNMR (100 MHz, CDCl3) δ 154.37, 144.69, 137.72, 131.68, 128.77, 128.41,123.00, 122.48, 120.53, 92.36, 85.82, 71.57, 66.03, 53.39, 44.85, 20.33.HRMS (ESI): exact mass calcd for C₁₈H₁₈N₂O [M+H]+, 279.149, found279.1513. As shown in FIG. 22, reverse phase HPLC was performed onRestek's Ultra IBD C18 (5 μm, 4.6×50 mm) using two Shimadzu LC-20ADpumps and a SPD-20A-vis detector set at 330 nm: Method A, 10%-40%acetonitrile in H₂O (v/v), flow rate at 1 mL/min over 20 min; method B,8%-40% methanol in H₂O (v/v), flow rate at 1 mL/min over 20 min.

Example 2

Synthesis of 3-fluoro-5-iodo-N,N-dimethylaniline (VMY-2-119)

The suspension of 1-fluoro-3-iodo-5-nitrobenzene (0.5 g, 1.87 mmol) andSnCl₂.2H₂O (1.5 g, 6.64 mmol) in EtOH (10 mL) was heated to reflux for1.5 h. The solvent was removed and the crude mixture was diluted withether, washed with 4 N NaOH and brine. The ether layer was separated anddried over Na₂SO₄, filtered, concentrated to yield amine compound as asolid (0.4 g, 91%). The crude product was used without purification.

A solution of above amine (0.4 g, 1.69 mmol) and iodomethane (0.719 g, 5mmol) in dimethylformamide (DMF; 10 mL) containing potassium carbonate(0.46 g, 3.38 mmol) was stirred for 48 h at room temperature. Water (10mL) was then added and the solution was extracted with ether threetimes. The organic extracts washed with water, brine, dried over Na₂SO₄,filtered and concentrated. The crude product was purified by columnchromatography to yield VMY-2-119 as a liquid (0.23 g, 52%). ¹H NMR (399MHz) δ 6.69-6.63 (m, 2H), 6.22 (dt, J=12.5, 2.3, 1H), 2.83 (s, 6H). ¹³CNMR (100 MHz) δ 163.33 (d, JF-C=245 Hz), 152.49 (d, J=11 Hz) 116.91 (d,J=2.4), 112.12 (d, J=24 Hz), 98.73 (d, J=26 Hz), 94.24 (d, J=11 Hz),40.21 (s, 3H).

Example 3

(S)-tert-butyl-2-((5-bromopyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-3)

Method A was used. Yield 55% (white solid). ¹H NMR (400 MHz, CDCl3) δ8.25-8.19 (m, 2H), 7.36 (s, 1H), 4.44 (d, J=5.3, 1H), 4.32-4.20 (m, 1H),4.06 (dd, J=2.8, 10.1, 1H), 3.81 (t, J=7.5, 2H), 2.36-2.14 (m, 2H), 1.36(s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 156.07, 155.41, 143.13, 136.65,124.02, 120.28, 79.76, 69.00, 59.92, 47.14, 28.37, 18.95.HRMS (ESI):exact mass calcd for C₁₄H₁₉BrN₂O₃ [M+H]+, 343.0657, found 343.0670.

Example 4

(S)-tert-butyl-2-((5-((trimethylsilyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-87)

Yield 62% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.04 (s, 2H), 7.08 (d,1H), 4.26 (s, 1H), 4.09 (s, 1H), 3.89 (s, 1H), 3.63 (s, 2H), 2.07 (d,2H), 1.18 (s, 9H), 0.16 (s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 156.18,154.54, 145.22, 138.43, 123.38, 120.54, 101.53, 98.11, 79.76, 68.91,60.24, 47.27, 28.59, 19.21, 0.28.

Example 5

(S)-tert-butyl2-((5-ethynylpyridin-3-yloxy)methyl)azetidine-1-carboxylate

¹H NMR (400 MHz, CDCl3) δ 8.36 (s, 2H), 7.35 (s, 1H), 4.65-4.46 (m, 1H),4.35 (s, 1H), 4.16 (dd, J=2.9, 10.1, 1H), 3.91 (t, J=7.6, 2H), 3.22 (s,1H), 2.50-2.19 (m, 2H), 1.45 (s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 156.12,155.50, 145.25, 138.47, 123.68, 80.42, 80.15, 79.78, 68.81, 60.00,47.13, 28.39, 19.03.

Example 6

(S)-tert-butyl-2-((5-(phenylethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-89)

Method B in scheme 1 was used. Yield 54% (liquid). ¹H NMR (400 MHz,CDCl3) δ 8.39 (s, 1H), 8.31 (s, 1H), 7.59-7.51 (m, 2H), 7.37 (ddd,J=1.5, 3.4, 5.9, 4H), 4.62-4.44 (m, 1H), 4.36 (s, 1H), 4.16 (dd, J=2.9,10.1, 1H), 3.90 (t, J=7.6, 2H), 2.51-2.19 (m, 2H), 1.44 (s, 9H). ¹³C NMR(100 MHz, CDCl3) δ 156.12, 154.48, 144.78, 137.91, 131.66, 128.78,128.41, 122.99, 122.46, 120.58, 92.44, 85.79, 79.75, 68.80, 60.04,47.00, 28.41, 19.06. HRMS (ESI): exact mass calcd for C₂₂H₂₄N₂O₃ [M+H]¹,365.1870, found 365.1870.

Example 7

(S)-3-(azetidin-2-ylmethoxy)-5-(phenylethynyl)pyridine (VMY-2-95

Method D was used. Yield 70% (liquid). ¹H NMR (400 MHz, CDCl3) δ 8.37(s, 1H), 8.31-8.25 (m, 1H), 7.60-7.49 (m, 2H), 7.41-7.29 (m, 4H),4.33-4.20 (m, 1H), 4.10-3.96 (m, 2H), 3.69 (q, J=7.9, 1H), 3.45 (td,J=4.8, 8.1, 1H), 2.45-2.19 (m, 3H). ¹³C NMR (100 MHz, CDCl3) δ 154.43,144.56, 137.76, 131.63, 128.75, 128.38, 122.90, 122.43, 120.46, 92.33,85.85, 72.77, 56.99, 44.20, 23.89. HRMS (ESI): exact mass calcd forC₁₇H₁₆N₂O [M+H]′, 265.1341, found 265.1339. Anal. Calcd for C₁₇H₁₆N₂O.

Example 8

(S)-tert-butyl-2-((5-((4-fluorophenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-97)

Method C in Scheme 1 was used. Yield 63% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.37 (d, J=1.0, 1H), 8.31 (d, J=2.8, 1H), 7.57-7.50 (m, 2H),7.40-7.35 (m, 1H), 7.12-7.03 (m, 2H), 4.58-4.50 (m, 1H), 4.36 (s, 1H),4.16 (dd, J=2.9, 10.1, 1H), 3.90 (t, J=7.6, 2H), 2.53-2.18 (m, 2H), 1.44(s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 162.72 (d, J_(F-C)=250 Hz, C),156.08, 154.47, 144.66, 137.94, 133.58 (d, J_(F-C)=8.4 Hz,), 122.93,120.38, 118.56 (d, J_(F-C)=3.6), 115.72 (d, J_(F-C)=22), 91.32, 85.50(d, J_(F-C)=1.3) 79.70, 68.76, 60.02, 47.13, 28.37, 19.0. HRMS (ESI):exact mass calcd for C₂₂H₂₃FN₂O3 [M+H]+, 383.1771, found 383.1767.

Example 9

(S)-3-(azetidin-2-ylmethoxy)-5-((4-fluorophenyl)ethynyl)pyridine(VMY-2-101)

Method D was used. Yield 77% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.27(s, 1H), 8.19 (d, J=2.8, 1H), 7.47-7.39 (m, 2H), 7.23 (dd, J=1.5, 2.7,1H), 7.00-6.93 (m, 2H), 4.20 (s, 1H), 4.04-3.90 (m, 2H), 3.62 (d, J=7.2,1H), 3.37 (s, 1H), 2.37-2.14 (m, 3H). ¹³C NMR (100 MHz, CDCl3) δ 162.73(d, J_(F-C)=249 Hz, C), 154.45, 144.54, 137.83 133.60 (d, J_(F-C)=8.5),122.88, 120.30, 118.58 (d, J_(F-C)=3.5), 115.73 (d, J_(F-C)=23 Hz),91.22, 85.58, 72.84, 57.01, 44.22, 23.92. HRMS (ESI): exact mass calcdfor C₁₇H₁₅FN₂O [M+H]+, 283.1247, found 283.1241. Anal. Calcd forC₁₇H₁₅FN₂O.0.1 H₂O.

Example 10

(S)-tert-butyl2-((5-((2-fluorophenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-103)

Method C in Scheme 1 was used. Yield 77% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.32 (d, J=4.1, 1H), 8.24 (dd, J=2.8, 5.7, 1H), 7.45 (d, J=5.4,1H), 7.29 (d, J=16.2, 2H), 7.06 (dd, J=9.0, 10.1, 2H), 4.45 (s, 1H),4.28 (s, 1H), 4.08 (dd, J=3.4, 6.6, 1H), 3.88-3.72 (m, 2H), 2.25 (m,2H), 1.35 (s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 162.61 (d, J_(F-C)=251 HzC), 156.07, 154.44, 144.74, 138.24, 133.42 (d, J_(F-C)=0.9), 130.56 (d,J_(F-C)=8.0), 124.03 (d, J_(F-C)=3.8), 122.97, 120.16, 115.56 (d,J_(F-C)=21 Hz), 111.11 (d, J_(F-C)=15 Hz), 90.71 (d, J_(F-C)=3.2 Hz),85.73, 79.70, 68.78, 60.02, 28.37, 47.12, 19.02. HRMS (ESI): exact masscalcd for C₂₂H₂₃FN₂O₃ [M+H]⁺, 383.1771, found 383.1784.

Example 11

(S)-3-(azetidin-2-ylmethoxy)-5-((2-fluorophenyl)ethynyl)pyridine(VMY-2-105)

Method D was used. Yield 81% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.31(d, J=1.0, 1H), 8.21 (dd, J=2.7, 0.9, 1H), 7.49-7.41 (m, 1H), 7.31-7.23(m, 2H), 7.11-7.00 (m, 2H), 4.29-4.14 (m, 1H), 4.05-3.91 (m, 2H), 3.63(q, J=7.9, 1H), 3.39 (td, J=7.8, 5.0, 1H), 2.41-2.11 (m, 3H). ¹³C NMR(100 MHz, CDCl3) δ 162.55 (d, J_(F-C)=251 Hz, C), 154.39, 144.49,138.10, 133.37 (d, J_(F-C)=1.0), 130.51 (d, J_(F-C)=8.0), 124.00 (d,J_(F-C)=3.7), 122.83, 120.03, 115.50 (d, J_(F-C)=21 Hz), 111.07 (d,J_(F-C)=16 Hz), 90.76 (d, J_(F-C)=3.3, 3H), 85.61, 72.76, 56.93, 44.15,23.82. HRMS (ESI): exact mass calcd for C₁₇H₁₅FN₂O [M+H]+, 283.1247,found 283.1243. Anal. Calcd for C₁₇H₁₅FN₂O.

Example 12

(S)-tert-butyl2-((5-((3-fluorophenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-107)

Method C in Scheme 1 was used. Yield 48% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.30 (s, 1H), 8.24 (d, J=2.6, 1H), 7.26 (ddd, J=12.6, 8.1, 1.1,3H), 7.20-7.12 (m, 1H), 7.05-6.94 (m, 1H), 4.45 (s, 1H), 4.29 (s, 1H),4.08 (dd, J=10.1, 2.8, 1H), 3.82 (t, J=7.5, 2H), 2.41-2.12 (m, 2H), 1.37(s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 162.34 (d, J_(F-C), 246 Hz), 156.13,154.51, 144.78, 138.25, 130.04 (d, J_(F-C)=8.6, 7H), 127.57 (dJ_(F-C)=3.1), 124.30 (d, J_(F-C)=9 Hz) 123.05, 120.10, 118.42 (d,J_(F-C)=23 Hz), 116.14 (d, J_(F-C)=21 Hz), 91.05 (d, J_(F-C)=3.4),86.65, 79.76, 68.82, 60.05, 47.11, 28.4, 19.05. HRMS (ESI): exact masscalcd for C₂₂H₂₃FN₂O₃ [M+H]⁺, 383.1771, found 383.1778.

Example 13

(S)-3-(azetidin-2-ylmethoxy)-5-((3-fluorophenyl)ethynyl)pyridine(VMY-2-109

Method D was used. Yield 78% (Liquid). ¹H NMR (399 MHz, CDCl3) δ 8.30(d, J=1.6, 1H), 8.22 (d, J=2.8, 1H), 7.30-7.23 (m, 3H), 7.20-7.13 (m,1H), 7.07-6.95 (m, 1H), 4.29-4.15 (m, 1H), 3.98 (qd, J=9.5, 5.5, 2H),3.65 (q, J=7.9, 1H), 3.46-3.35 (m, 1H), 2.42-2.13 (m, 3H). ¹³C NMR (100MHz, CDCl3) δ 162.33 (d, J_(F-C)=246 Hz) 154.46, 144.62, 138.09, 130.01(d, J_(F-C)=9 Hz), 127.56 (d, J_(F-C)=3.1 Hz), 124.29 (d, J_(F-C)=9 Hz),123.00, 120.02, 118.42 (d, J_(F-C)=23 Hz), 116.12 (d, J_(F-C)=21 Hz),116.02, 90.96 (d, J_(F-C)=3.4), 72.84, 57.00, 44.22, 23.92. HRMS (ESI):exact mass calcd for C₁₇H₁₃FN₂O [M+H]⁺, 283.1247, found 283.1236. Anal.Calcd for C₁₇H₁₃FN₂O.

Example 14

(S)-tert-butyl-2-((5-((3,5-difluorophenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-111)

Method B in Scheme 1 was used. Yield 60% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.30 (s, 1H), 8.26 (d, J=2.8, 1H), 7.30 (s, 1H), 7.02-6.93 (m,2H), 6.76 (tt, J=8.9, 2.2, 1H), 4.54-4.40 (m, 1H), 4.29 (s, 1H),4.15-4.03 (m, 1H), 3.82 (t, J=7.6, 2H), 2.40-2.12 (m, 2H), 1.36 (s, 9H).¹³C NMR (100 MHz, CDCl3) δ 163.89 (d, J_(F-C)=13 Hz), 161.41 (d,J_(F-C)=13 Hz), 156.10, 154.48, 144.76, 138.57, 125.11 (t), 123.03,119.57, 114.68 (d, J_(F-C)=7.7), 114.49 (d, J_(F-C)=7.6), 104.90 (t),89.88, 87.65, 79.73, 68.80, 60.01, 47.11, 28.36, 18.99. HRMS (ESI):exact mass calcd for C₂₂H₂₂F₂N₂O3 [M+H]+, 401.1677, found 401.1692.

Example 15

(S)-3-(azetidin-2-ylmethoxy)-5-((3,5-difluorophenyl)ethynyl)pyridine(VMY-2-113)

Method B in Scheme 1 was used. Yield 79% (liquid). ¹H NMR (400 MHz,CDCl3) δ 8.29 (d, J=1.6, 1H), 8.24 (d, J=2.8, 1H), 7.25 (dd, J=1.7, 2.8,1H), 7.03-6.94 (m, 2H), 6.77 (tt, J=2.3, 8.9, 1H), 4.23 (s, 1H), 3.99(qd, J=5.5, 9.5, 2H), 3.66 (d, J=7.6, 1H), 3.39 (s, 1H), 2.48-1.79 (m,3H). ¹³C NMR (100 MHz) δ 163.91 (d, J_(F-C)=13 Hz), 161.43 (d,J_(F-C)=13 Hz), 154.47, 144.63, 138.43, 125.13 (t), 123.02, 119.51,114.71 (d, =8 Hz), 114.51 (d, =7.7), 104.92 (t), 89.83 (t, =3.9), 87.70,72.90, 56.99, 44.22, 23.9. HRMS (ESI): exact mass calcd for C₁₇H₁₅FN₂O[M+H]+, 301.1152, found 301.1155. Anal. Calcd for C₁₇H₁₄F₂N₂O.0.06 H₂O.

Example 16

(S)-tert-butyl-2-((5-((3-(trifluoromethyl)phenyl)ethynyl)pyridin-3yloxy)methyl)azetidine-1-carboxylate (VMY-2-115)

Method B in Scheme 1 was used. Yield 50% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.39 (d, J=1.4, 1H), 8.33 (d, J=2.8, 1H), 7.80 (s, 1H), 7.70(d, J=7.8, 1H), 7.61 (d, J=7.9, 1H), 7.50 (t, J=7.8, 1H), 7.40 (d,J=1.8, 1H), 4.53 (dd, J=5.4, 8.1, 1H), 4.37 (s, 1H), 4.22-4.10 (m, 1H),3.90 (t, J=7.6, 2H), 2.49-2.17 (m, 2H), 1.55-1.32 (m, 9H). ¹³C NMR (100MHz, CDCl3) δ 156.03, 154.48, 144.66, 138.40, 134.63, 130.90, 128.93,128.34, 125.17, 123.40, 122.92, 119.79, 90.59, 87.27, 79.59, 68.76,60.01, 47.13, 28.28, 18.92. HRMS (ESI): exact mass calcd forC₂₃H₂₃F₃N₂O₃ [M+H]+, 433.1739, found 433.1749.

Example 17

(S)-3-(azetidin-2-ylmethoxy)-5-((3-(trifluoromethyl)phenyl)ethynyl)pyridine(VMY-2-117)

Method D was used. Yield 78% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.31(d, J=1.6, 1H), 8.24 (d, J=2.8, 1H), 7.74 (s, 1H), 7.64 (d, J=7.7, 1H),7.55 (d, J=7.9, 1H), 7.43 (t, J=7.8, 1H), 7.27 (dd, J=1.7, 2.8, 1H),4.23 (td, J=7.5, 12.3, 1H), 3.99 (qd, J=5.5, 9.5, 2H), 3.66 (dd, J=8.1,15.9, 1H), 3.40 (dt, J=4.9, 7.6, 1H), 2.52-1.90 (m, 3H). ¹³C NMR (100MHz, CDCl3) δ 154.48, 144.63, 138.27, 134.69 (d, J=1.1), 131.06 (q),128.96, 128.45 (d, J=3 Hz), 125.27 (d, J=3.7), 123.45, 123.01, 119.82,90.62, 87.32, 72.86, 57.01, 44.22, 23.90. HRMS (ESI): exact mass calcdfor C₁₈H₁₅F₃N₂O [M+H]+, 333.1215, found 333.1216. Anal. Calcd forC₁₈H₁₅F₃N₂O.

Example 18

(S)-tert-butyl-2-((5-(m-tolylethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-121)

Method B was used. Yield 56% (liquid). ¹H NMR (400 MHz, CDCl3) δ 8.38(d, J=1.6, 1H), 8.30 (d, J=2.8, 1H), 7.40-7.33 (m, 3H), 7.29-7.24 (m,1H), 7.19 (d, J=7.6, 1H), 4.59-4.48 (m, 1H), 4.36 (s, 1H), 4.22-4.12 (m,1H), 3.90 (t, J=7.6, 2H), 2.43-2.33 (s, 3H), 1.44 (s, 9H). ¹³C NMR (100MHz, CDCl3) δ 156.09, 154.47, 144.75, 138.09, 137.81, 132.22, 129.69,128.73, 128.30, 122.95, 122.25, 120.67, 92.67, 85.45, 79.71, 68.78,60.05, 47.06, 28.40, 21.18, 19.05. HRMS (ESI): exact mass calcd forC₂₃H₂₆N₂O₃ [M+H]+, 379.2022, found 379.2031.

Example 19

(S)-3-(azetidin-2-ylmethoxy)-5-(m-tolylethynyl)pyridine (VMY-2-123)

Method D was used. Yield 68% (Liquid). ¹H NMR (400 MHz) δ 8.26 (t,J=2.9, 1H), 8.17 (d, J=2.8, 1H), 7.30-7.21 (m, 3H), 7.15 (t, J=7.6, 1H),7.10-7.05 (m, 1H), 4.24-4.12 (m, 1H), 3.94 (qd, J=9.5, 5.5, 2H), 3.60(q, J=8.1, 1H), 3.36 (td, J=8.2, 4.4, 1H), 2.44 (s, 1H), 2.35-2.09 (m,5H). ¹³C NMR (100 MHz,) δ 154.43, 144.57, 138.09, 137.67, 132.20,129.68, 128.73, 128.29, 122.90, 122.23, 120.59, 92.59, 85.49, 72.72,57.01, 44.19, 23.88, 21.17. HRMS (ESI): exact mass calcd for C₁₈H₁₈N₂O[M+H]+, 279.1497, found 279.1512. Anal. Calcd for C₁₈H₁₈N₂O.0.06 CH₂Cl₂.

Example 20

(S)-tert-butyl2-((5-((3-chlorophenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-125)

Method B was used. Yield 65% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.29(d, J=1.5, 1H), 8.24 (d, J=2.8, 1H), 7.44 (dd, J=2.5, 0.9, 1H), 7.33(dt, J=7.4, 1.5, 1H), 7.28 (ddd, J=3.4, 2.4, 1.5, 1H), 7.25 (dd, J=2.0,1.4, 1H), 7.24-7.18 (m, 1H), 4.56-4.38 (m, 1H), 4.28 (s, 1H), 4.08 (dd,J=10.1, 2.9, 1H), 3.81 (t, J=7.6, 2H), 2.37-2.12 (m, 2H), 1.35 (s, 9H).¹³C NMR (100 MHz, CDCl3) δ 156.08, 154.46, 144.72, 138.24, 134.23,131.44, 129.74, 129.63, 128.99, 124.16, 122.99, 120.02, 90.86, 86.93,79.71, 68.78, 60.01, 47.01, 28.38, 19.02. HRMS (ESI): exact mass calcdfor C₂₂H₂₃ClN₂O₃ [M+H]+, 399.1475, found 399.1456.

Example 21

(S)-3-(azetidin-2-ylmethoxy)-5-((3-chlorophenyl)ethynyl)pyridine(VMY-2-127)

Method D was used. Yield 72% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.27(d, J=1.6, 1H), 8.20 (d, J=2.8, 1H), 7.44 (t, J=1.7, 1H), 7.33 (dt,J=7.4, 1.4, 1H), 7.27-7.17 (m, 3H), 4.26-4.14 (m, 1H), 4.04-3.89 (m,2H), 3.62 (q, J=8.1, 1H), 3.43-3.32 (m, 1H), 2.38-2.11 (m, 3H). ¹³C NMR(100 MHz, CDCl3) δ 154.44, 144.57, 138.09, 134.23, 131.46, 129.75,129.62, 128.99, 124.16, 122.95, 119.96, 90.78, 86.96, 72.81, 56.98,44.20, 23.89. HRMS (ESI): exact mass calcd for C₁₇H₁₅ClN₂O [M+H]+,299.0951, found 299.0965. Anal. Calcd for C₁₇H₁₅ClN₂O.0.06 H₂O.

Example 22

(S)-tert-butyl2-((5-((3-(dimethylamino)-5-fluorophenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate (VMY-2-129)

Method B in Scheme 1 was used. Yield 14% (Liquid). ¹H NMR (400 MHz) δ8.35 (s, 1H), 8.27 (s, 1H), 7.35 (s, 1H), 6.60 (s, 1H), 6.54 (d, J=8.7,1H), 6.37 (dt, J=12.4, 2.2, 1H), 4.49 (s, 1H), 4.33 (s, 1H), 4.19-4.08(m, 1H), 3.87 (t, J=7.6, 2H), 2.94 (d, J=9.7, 6H), 2.43-2.20 (m, 2H),1.40 (s, 9H). ¹³C NMR (100 MHz) δ 163.55 (d, J_(F-C)=241 Hz), 156.14,154.48, 151.68 (d, J_(F-C)=12 Hz), 144.80, 138.00, 123.86 (d, J_(F-C)=12Hz), 123.06, 111.13 (d, J_(F-C)=2.0), 106.45 (d, J_(F-C)=76 Hz), 100.1(d, J_(F-C)=26 Hz) 92.42 (d, =4.2), 85.17, 79.79, 68.81 (d, J_(F-C)=3Hz), 60.03, 47.01, 40.29, 28.42, 19.07.

Example 23

(S)-3-((5-(azetidin-2-ylmethoxy)pyridin-3-yl)ethynyl)-5-fluoro-N,N-dimethylaniline(VMY-2-131)

Method D was used. Yield 72% (Liquid). ¹H NMR (400 MHz) δ 8.29 (d,J=1.6, 1H), 8.21 (t, J=4.5, 1H), 7.26 (dd, J=2.8, 1.7, 1H), 6.58-6.46(m, 2H), 6.32 (dt, J=12.5, 2.3, 1H), 4.21 (d, J=15.8, 1H), 3.99 (qd,J=9.5, 5.5, 2H), 3.65 (q, J=7.9, 1H), 3.41 (dd, J=12.0, 8.3, 1H),2.93-2.85 (s, 6H), 2.40-2.13 (m, 2H), 2.00 (d, J=19.3, 1H). ¹³C NMR (100MHz) δ 206.86 (Acetone, C═O), 163.55 (d, J_(F-C)=241 Hz), 154.44, 151.68(d, J_(F-C)=11 Hz), 144.69, 137.89, 123.87 (d, J_(F-C)=12 Hz), 123.02,120.36, 111.12 (d, J_(F-C)=2.0, 1H), 105.97 (d, J_(F-C)=24 Hz), 100.10(d, J_(F-C)=26 Hz), 92.33 (d, J_(F-C)=4 Hz), 85.22, 72.73, 57.06, 44.23,40.29, 30.88 (Acetone), 23.90. HRMS (ESI): exact mass calcd forC₁₉H₂₀FN₃O [M+H]+, 326.1669, found 326.1668.

As shown in FIG. 22, reverse phase HPLC was performed on Restek's UltraIBD C18 (5 μm, 4.6×50 mm) using two Shimadzu LC-20AD pumps and aSPD-20A-vis detector set at 330 nm: Method A, 10%-40% acetonitrile inH₂O (v/v), flow rate at 1 mL/min over 20 min; method B, 8%-40% methanolin H₂O (v/v), flow rate at 1 mL/min over 20 min.

Example 24

(S)-tert-butyl-2-((5-((3-fluoro-5-methylphenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate (VMY-2-133)

Method B in Scheme 1 was use. Yield 65% (liquid). ¹H NMR (400 MHz,CDCl3) δ 8.28 (d, J=1.5, 1H), 8.22 (d, J=2.8, 1H), 7.28 (dd, J=2.6, 1.7,1H), 7.06 (d, J=0.5, 1H), 6.94 (dd, J=9.1, 0.5, 1H), 6.80 (d, J=9.5,1H), 4.51 4.39 (m, 1H), 4.27 (s, 1H), 4.12-4.00 (m, 1H), 3.81 (t, J=7.6,2H), 2.35-2.09 (m, 5H), 1.35 (s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 162.29(d, J_(F-C)=245 Hz, C), 156.10, 151.47, 144.75, 140.56 (d, J_(F-C)=8Hz), 138.13, 128.19 (d, J_(F-C)=2.8, 3H), 123.75 (d, J_(F-C)=11 Hz),123.00, 120.19, 116.86 (d, J_(F-C)=21 Hz), 115.40 (d, J_(F-C)=23 Hz),91.33 (d, J_(F-C) 3.7), 86.18, 79.73, 68.81, 60.04, 47.20, 28.38, 21.13,19.03. HRMS (ESI): exact mass calcd for C₂₃H₂₅FN₂O₃ [M+H]+, 397.1927,found 397.1929.

Example 25

(S)-3-(azetidin-2-ylmethoxy)-5-((3-fluoro-5-methylphenyl)ethynyl)pyridine(VMY-2-135)

Method D was used. Yield 73% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.33(d, J=1.6, 1H), 8.29-8.24 (m, 1H), 7.29 (dd, J=2.8, 1.7, 1H), 7.12 (dt,J=2.1, 0.7, 1H), 7.05-6.98 (m, 1H), 6.87 (dddd, J=9.6, 2.3, 1.4, 0.7,1H), 4.28 (tt, J=12.3, 6.2, 1H), 4.03 (qd, J=9.5, 5.5, 2H), 3.70 (dd,J=15.9, 8.2, 1H), 3.45 (ddd, J=7.6, 7.1, 4.4, 1H), 2.43-2.34 (m, 2H),2.33 (t, J=1.5, 3H), 2.25 (ddd, J=16.4, 11.1, 8.2, 1H). ¹³C NMR (100MHz, CDCl3) δ 162.31 (d, J_(F-C)=246.3), 154.44, 144.65, 140.57 (d,J_(F-C)=8.6), 138.01, 128.20 (d, J_(F-C)=2.7), 123.77 (d, J_(F-C)=10.3),122.98, 120.13, 116.88 (d, J_(F-C)=21.1), 115.44 (d, J_(F-C)=23.1),91.26 (d, J_(F-C)=3.7), 86.23, 72.78, 57.04, 44.22, 23.88, 21.12. HRMS(ESI): exact mass calcd for C₁₈H₁₇FN₂O [M+H]+, 297.1403, found 297.1403.Anal. Calcd for C₁₈H₁₇FN₂O.0.04 H₂O.

Example 26

(S)-tert-butyl-2-((5-((3-fluoro-5-methoxyphenyl)ethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate (VMY-2-137)

Method B in Scheme 1 was used. Yield 37% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.27 (s, 1H), 8.21 (d, J=2.7, 1H), 7.31-7.24 (m, 1H), 6.74(ddd, J=1.2, 2.1, 9.9, 2H), 6.54 (dt, J=2.3, 10.6, 1H), 4.47-4.38 (m,1H), 4.25 (s, 1H), 4.09-4.02 (m, 1H), 3.79 (t, J=7.6, 2H), 3.72 (s, 3H),2.32-2.13 (m, 2H), 1.32 (s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 163.14 (d,J_(F-C)=245.8), 160.74 (d, J_(F-C)=12.1), 156.13, 154.49, 144.80,138.27, 124.34 (d, J_(F-C)=12.1), 123.07, 120.06, 112.82 (d,J_(F-C)=2.9), 110.94 (d, J_(F-C)=23.5), 103.22 (d, J_(F-C)=25.0), 91.24,86.33, 79.78, 68.82, 60.03, 55.66, 47.36, 28.40, 19.05. HRMS (ESI):exact mass calcd for C₂₃H₂₅FN₂O₄ [M+H]+, 413.1877, found 413.1887.

Example 27

(S)-3-(azetidin-2-ylmethoxy)-5-((3-fluoro-5-methoxyphenyl)ethynyl)pyridine(VMY-2-139)

Method D was used. Yield 66% (Liquid). ¹H NMR (400 MHz, CDCl3) δ 8.29(d, J=1.6, 1H), 8.22 (d, J=2.8, 1H), 7.25 (dd, J=2.8, 1.7, 1H),6.83-6.72 (m, 2H), 6.57 (dt, J=10.6, 2.3, 1H), 4.22 (s, 1H), 4.06-3.92(m, 2H), 3.75 (s, 3H), 3.64 (s, 1H), 3.39 (s, 1H), 2.54-1.91 (m, 3H).¹³C NMR (100 MHz, CDCl3) δ 163.16 (d, =245.6), 160.74 (d, J_(F-C)=12.1),154.48, 144.68, 138.14, 124.37 (d, J_(F-C)=12.1), 123.03, 119.99, 112.84(d, J_(F-C)=2.9), 110.97 (d, J_(F-C)=23.6), 103.21 (d, J_(F-C)=25.0),91.14 (d, J_(F-C)=4.2), 86.40, 72.89, 57.03, 55.68, 44.24, 23.95. HRMS(ESI): exact mass calcd for C₁₈H₁₇FN₂O₂ [M+H]+, 313.1352, found313.1358. Anal. Calcd for C₁₈H₁₇FN₂O₂.0.04 H₂O.

Example 28

3-bromo-5-(cyclobutylmethoxy)pyridine (VMY-2-157)

Method A was used. Yield 69% (Liquid). ¹H NMR (399 MHz, CDCl3) δ 8.18(dd, J=11.0, 2.1, 2H), 7.28 (dd, J=2.5, 1.9, 1H), 3.88 (dd, J=6.4, 3.4,2H), 2.78-2.64 (m, 1H), 2.14-2.02 (m, 2H), 1.99-1.74 (m, 4H). ¹³C NMR(100 MHz, CDCl3) δ 155.44, 142.34, 136.33, 123.39, 120.07, 72.30, 34.16,24.53, 18.40. HRMS (ESI): exact mass calcd for C₁₀H₁₂BrNO [M+H]+,242.0181; found 242.0181.

Example 29

3-(cyclobutylmethoxy)-5-(phenylethynyl)pyridine (VMY-2-161)

Method B in Scheme 1 was used. Yield 65% (Liquid). ¹H NMR (400 MHz,CDCl3) δ 8.36 (d, J=1.5, 1H), 8.26 (d, J=2.8, 1H), 7.60-7.50 (m, 2H),7.40-7.33 (m, 3H), 7.34-7.28 (m, 1H), 3.98 (d, J=6.6, 2H), 2.88-2.73 (m,1H), 2.23-2.10 (m, 2H), 2.07-1.82 (m, 4H). ¹³C NMR (100 MHz, CDCl3) δ154.72, 144.33, 137.86, 131.67, 128.75, 128.41, 122.86, 122.54, 120.47,92.24, 85.96, 72.42, 34.41, 24.76, 18.54. HRMS (ESI): exact mass calcdfor C₁₈H₁₇NO [M+H]+, 264.1388, found 264.1396. Anal. Calcd for C₁₈H₁₇NO.

Example 30

tert-butyl-2-(5-bromopyridin-3-yloxy)ethyl(propyl)carbamate (VMY-2-163)

Method A in FIG. 3 was used. Yield 62% (Liquid). ¹H NMR (400 MHz, CDCl3)δ 8.21 (s, 1H), 8.15 (d, J=2.5, 1H), 7.29 (s, 1H), 4.06 (s, 2H), 3.51(s, 2H), 3.16 (s, 2H), 1.56-1.43 (m, 2H), 1.39 (s, 9H), 0.82 (t, J=7.4,3H). ¹³C NMR (100 MHz, CDCl3) δ 155.05, 142.72, 136.36, 123.45, 120.14,79.37, 67.00, 50.40, 46.62, 28.22, 21.84, 11.04. HRMS (ESI): exact masscalcd for C₁₃H₂₃BrN₂O₃ [M+H]+, 359.0970; found 359.0978.

Example 31

tert-butyl-2-(5-(phenylethynyl)pyridin-3-yloxy)ethyl(propyl)carbamate(VMY-2-171)

Method B in Scheme 1 was used. Yield 53% (solid). ¹H NMR (399 MHz,CDCl3) δ 8.37 (s, 1H), 8.25 (d, J=2.8, 1H), 7.59-7.49 (m, 2H), 7.37-7.28(m, 4H), 4.14 (s, 2H), 3.59 (s, 2H), 3.25 (s, 2H), 1.64-1.51 (m, 2H),1.47 (s, 9H), 0.89 (t, J=7.4, 3H). ¹³C NMR (100 MHz, CDCl3) δ 155.62,154.21, 144.55, 137.67, 131.60, 128.73, 128.36, 122.59, 122.41, 120.54,92.43, 85.79, 79.55, 66.95, 50.56, 46.85, 28.37, 21.95, 11.16. HRMS(ESI): exact mass calcd for C₂₃H₂₈N₂O₃ [M+H]+, 381.2187; found 381.2169.

Example 32

N-(2-(5-(phenylethynyl)pyridin-3-yloxy)ethyl)propan-1-amine (VMY-2-177)

Method D was used. Yield 87% (solid). ¹H NMR (400 MHz, CDCl3) δ 8.29 (d,J=1.5, 1H), 8.19 (d, J=2.8, 1H), 7.50-7.41 (m, 2H), 7.33-7.26 (m, 3H),7.26-7.22 (m, 1H), 4.05 (t, J=5.2, 2H), 2.96 (t, J=5.2, 2H), 2.58 (t,J=7.2, 2H), 1.47 (td, J=14.6, 7.3, 3H), 0.87 (t, J=7.4, 3H). ¹³C NMR(100 MHz, CDCl3) δ 154.38, 144.64, 137.70, 131.65, 128.77, 128.39,122.83, 122.45, 120.53, 92.37, 85.83, 68.05, 51.72, 48.51, 23.18, 11.70.HRMS (ESI): exact mass calcd for C₁₈H₂₀N₂O [M+H]+, 281.1654, found281.1645. Anal. Calcd for C₁₈H₂₀N₂O.

Example 33

(R)-tert-butyl 2-((5-bromopyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-181)

Method A in FIG. 3 was used. Yield 30% (solid). ¹H NMR (400 MHz, CDCl3)δ 8.20 (d, J=2.4, 2H), 7.35 (t, J=1.9, 1H), 4.48-4.40 (m, 1H), 4.26 (s,1H), 4.05 (dd, J=10.1, 2.7, 1H), 3.86-3.76 (m, 2H), 2.70-2.01 (m, 2H),1.35 (s, 9H). ¹³C NMR (100 MHz, CDCl3) δ 156.06, 155.41, 143.10, 136.62,124.04, 120.28, 79.75, 69.00, 59.91, 47.08, 28.36, 18.94. HRMS (ESI):exact mass calcd for C₁₄H₁₉BrN₂O₃ [M+H]+, 343.0657, found 343.0680.

Example 34

(R)-tert-butyl-2-((5-(phenylethynyl)pyridin-3-yloxy)methyl)azetidine-1-carboxylate(VMY-2-189)

Method B in Scheme 1 was used. Yield 74%. ¹H NMR (400 MHz, CDCl3) δ 8.31(s, 1H), 8.23 (s, 1H), 7.52-7.41 (m, 2H), 7.36-7.24 (m, 4H), 4.45 (d,J=5.4, 1H), 4.28 (s, 1H), 4.08 (dd, J=10.2, 2.6, 1H), 3.82 (t, J=7.6,2H), 2.28 (ddd, J=25.0, 14.3, 7.1, 2H), 1.36 (d, J=0.5, 9H). ¹³C NMR(100 MHz, CDCl3) δ 156.12, 154.50, 144.75, 137.86, 131.66, 128.78,128.40, 123.04, 122.46, 120.63, 92.46, 85.77, 79.76, 68.81, 47.01,60.05, 28.41, 19.06. HRMS (ESI): exact mass calcd for C₂₂H₂₄N₂O₃ [M+H]+,365.1865, found 343.1893.

Example 35

(R)-3-(azetidin-2-ylmethoxy)-5-(phenylethynyl)pyridine (VMY-2-191)

Method D was used. Yield 75% (Liquid). ¹H NMR (399 MHz, CDCl3) δ 8.28(s, 1H), 8.19 (d, J=2.8, 1H), 7.51-7.42 (m, 2H), 7.27 (ddd, J=10.0, 6.0,0.8, 4H), 4.26-4.14 (m, 1H), 3.96 (qd, J=9.5, 5.7, 2H), 3.62 (q, J=8.0,1H), 3.38 (td, J=8.1, 4.4, 1H), 2.38-2.12 (m, 3H). ¹³C NMR (100 MHz,CDCl3) δ 154.45, 144.61, 137.76, 131.66, 128.77, 128.40, 122.96, 122.46,120.51, 92.35, 85.84, 72.78, 57.03, 44.22, 23.91. HRMS (ESI): exact masscalcd for C₁₇H₁₆N₂O [M+H]+, 265.1341, found 265.1349. Anal. Calcd forC₁₇H₁₆N₂O.0.6 H₂O.

Example 36 (S)-3-(azetidin-2-ylmethoxy)-5-phenylpyridine (VMY-2-203)

This compound was prepared according to the method shown in FIG. 3. To asolution of(S)-tert-butyl-2-((5-bromopyridin-3-yloxy)methyl)azetidine-1-carboxylate(1, 0.58 mmol) in 9 mL of toluene and 3 mL of ethanol was added phenylboronic acid (0.69 mmol) followed by 2 mL of 2 M Na₂CO₃ andtetrakis(triphenylphosphine)palladium (0) (0.03 mmol). The reaction wasstirred for 12 h at 90° C. under nitrogen. The reaction was cooled toroom temperature, diluted with water and extracted three times withethyl acetate. The combined organic layers dried over anhydrous sodiumsulfate, filtered and concentrated in vacuo to afford a crude product,which was subsequently purified by column chromatography. The resultingpure compound was subjected to Boc deprotection followed by purificationof column chromatography to yield a final pure compound VMY-2-203 as aliquid (70 mg, 71%). ¹H NMR (400 MHz, CDCl₃): δ 8.37 (d, J=1.7, 1H),8.22 (d, J=2.7, 1H), 7.49 (dd, J=5.1, 3.8, 2H), 7.38 (t, J=7.6, 2H),7.31 (ddd, J=7.7, 4.1, 0.5, 2H), 4.31-4.14 (m, 1H), 4.02 (qd, J=9.5,5.6, 2H), 3.63 (q, 0.1=8.0, 1H), 3.39 (td, 0.1=8.2, 4.4, 1H), 2.39-2.13(m, 3H). ¹³C NMR (100 MHz, CDCl₃) δ 155.1, 140.7, 137.6, 137.2, 136.5,128.9, 128.1, 127.1, 119.7, 72.8, 57.1, 44.2, 24.0.HRMS (ESI): exactmass calcd for C₁₅H₁₆N₂O [M+H]⁺, 241.1341, found. 241.1348 Anal. Calcdfor C₁₅H₁₆N₂O.0.6H₂O. [α]_(D) ^(25.5)=−4.69 (c=1.4, CHCl₃).

Example 37

The in vitro binding affinities of the new ligands were measured fordefined receptor subtypes (α2β2, α2β4, α3β2, α3β4, α4β2, α4β4, α6β2,α6β4, and α7) expressed in stably transfected cell lines.[³H]Epibatidine ([³H]EB) binds to the agonist recognition site of all ofthe defined receptor subtypes with high affinities. Rat forebrainhomogenates were included to allow comparison between the heterologousand native α4β2 and α7 nAChRs. See FIG. 18 for binding affinity values(K_(i)) of the ligands at the three major nAChR subtypes (α3β4, α 4β32,and α7).

TABLE 1 In Vitro Pharmacological Properties of Nicotinic Ligands K_(i)(nM) IC_(50(10′)) (nM) Compound α3β4 α4β2 α7 α3β4 α4β2 Sazetidine-A1,900 0.062 1,600 >10,000 7.5 VMY-2-95 650 0.049 580 16 VMY-2-101 6500.083 2,000 8 VMY-2-105 580 0.072 1,100 20 VMY-2-109 520 0.032 720 13VMY-2-113 1,400 0.05 1,800 VMY-2-117 1,400 0.26 1,300 VMY-2-123 4,0000.046 200 VMY-2-127 3,400 0.093 480 VMY-2-131 1,000 0.031 250 VMY-2-1351,100 0.043 590 VMY-2-139 1,300 0.076 1,500 VMY-2-161 1,300,000 160,000410,000 >100,000 35,000 VMY-2-177 850,000 1,100 48,000 12,000 15,000VMY-2-191 1,700 0.11 1,600 21,000 16,000 VMY-2-205 40,000 1.2 7,70022,000 370

Example 38

Functional properties of the new ligands were determined by ⁸⁶Rb¹ effluxassays in cells expressing α3β4 and α4β2 nAChR subtypes. Functionalactivity of each new ligand was measured in terms of its agonism,antagonism and desensitization ability.

Agonist activity of each of the ligands was tested at eightconcentrations. The responses were compared to that stimulated by 100 μM(−)-nicotine, a near maximally effective concentration. The fullconcentration-effect curves generated potency (EC₅₀) and efficacy(E_(max)) of each ligand. The antagonist activity of each new compoundwas determined by applying the compound to cells simultaneously with 100μM (−)-nicotine.

Each compound was tested for antagonist activity at eightconcentrations. The potency (IC_(50(0′))) of each ligand as anantagonist was derived from the full concentration-effect curves.

The desensitization potency of each ligand was determined bypre-treating cells with the test compound for 10 minutes before 100 μM(−)-nicotine was applied. The potency of a compound to desensitize thereceptor after a 10 minute exposure (IC_(50(10′))) was obtained withfull concentration-effect curves using at least eight concentrations ofthe ligand. Though the ⁸⁶Rb⁺ efflux assays were the main methodologyused to determine functional properties, whole-cell current measurementswere also used to verify the key experiments. See Table 1 for potency ofthe compounds to desensitize the two major receptor (IC_(50(10′)))subtypes, α3β34 and α4β2.

Example 39

Preliminary studies of VMY-2-95 in animal models indicate that thecompound may be effective in reducing nicotine and ethanolself-administration. Previous studies have shown that Sazetidine-A, aselective α4β2 nicotinic receptor-desensitizing agent, was effective atreducing nicotine self-administration in rats, with lowest effectivedose of 3 mg/kg administered subcutaneously (sc). In this example,VMY-2-95 was assessed for efficacy in reducing nicotineself-administration in young adult female Sprague-Dawley rats (N=15).Before the start of nicotine self-administration sessions, all animalswere trained to lever press in a standard dual-lever operant chamber forfood reinforcement. After food training, the animals were surgicallyimplanted with jugular catheters. Animals had an initial ten session ofnicotine (0.03 mg/kg FRI) self-administration prior to VMY-2-95delivery. VMY-2-95 was delivered with acute sc injections 10 minutesbefore the start of 45 minute nicotine sessions in a repeated measurescounterbalanced design at one of three doses: 0.3, 1, and 3 mk/kg, orsaline vehicle. Alpha of p<0.05 (two-tailed) was used as the thresholdfor significance. Results are shown in FIG. 5 and FIG. 6.

The 3 mg/kg dose significantly reduced nicotine self-administration inthe rats relative to the saline vehicle (p<0.025); however, the lowerdoses studied did not produce significant decreases. VMY-2-95 showedsimilar potency as Sazetidine-A. The effect of VMY-2-95 appeared tobecome more pronounced during the second phase (see FIG. 5).

Example 40 Computational Studies.

To study (S)-(−)-nicotine and (R)-(−)-deschloroepibatidine binding withthe α4β2 nAChR in atomic detail, a structural model of the LSD of humanα4β2 nAChR was built by using the Homology Model software MODELLERX9.10.The reference template is the x-ray crystal structure of the AChBP (PDBentry of 1UW6) and the rat α4β2 x-ray structure (PDB: 1OLE). Multiplesequence alignment was generated by Psi-BLAST and ClusterW. Uponconstruction of the model, appropriate ionization states weremaintained, the side chains were relaxed to remove possible side chainatom contacts with the neighboring residues, different rotamer states ofthe residue were assigned, and then local side chain atom dynamicsfollowed by minimization were performed. Minimization and Moleculardynamics simulations were carried out using the SANDER module of AMBER10.0 with default parameters. The homology modeled structure wasvalidated with PROCHECK and WHATIF program.

Molecular docking was carried out using SurFlexDock Module of Sybyl-X(Tripos Inc. St. Louis, USA). However, to be consistent with thenicotine structural conformation (PDB: 1UW6), a manual interventionfollowed by constrained molecular dynamics simulations were carried out.This procedure was applied to VMY-2-95, Varinicline and Sazetidine A.The α4β2 nAChR compound complexes were refined by molecular dynamicssimulation using the Amber 10.0 with the PARM98 force-field parameter.The charge and force field parameters of the compounds were obtainedusing the most recent Antechamber module in the Amber 10.0 (4), wherecompounds were minimized at the MP2/6-31G* level using Gaussian 09. TheSHAKE algorithm was used to keep all bonds involving hydrogen atomsrigid. Weak coupling temperature and pressure coupling algorithms wereused to maintain constant temperature and pressure, respectively.Electrostatic interactions were calculated with the Ewald particle meshmethod with a dielectric constant at 1R_(ij) and a nonbonded cutoff of12 Å for the real part of electrostatic interactions and for van derWaals interactions. The total charge of the system was neutralized byaddition of a chloride ion. The system was solvated in a 12 Å cubic boxof water where the TIP3P model was used. 5000 steps of minimization ofthe system were performed in which the α4β2 nAChR was constrained by aforce constant of 75 kcal/mol/Å. After minimization, a 20 ps simulationwas used to gradually raise the temperature of the system to 298 K whilethe complex was constrained by a force constant of 20 kcal/mol/Å.Another 20 ps equilibration run was used where only the backbone atomsof the complex were constrained by a force constant of 5 kcal/mol/Å.Final production run of 200 ps was performed with no constraints. Whenapplying constraints, the initial complex structure was used as areference structure. The PME method was used and the time step was 5 fs,and a neighboring pairs list was updated every 25 steps.

Binding Affinities for nAChR Subtypes.

The binding affinities of all compounds synthesized for the receptorsubtypes were examined in binding competition studies against[³H]-epibatidine. K_(i) values of these compounds at seven definedsubtypes of rat nAChRs and native nAChRs from rat forebrain are providedin FIGS. 14 and 15.

All compounds in series 1 exhibited high affinity for the rat α4β2 nAChRsubtype with K_(i) values ranging from 0.031 nM (VMY-2-131) to 0.26 nM(VMY-2-117). These compounds also showed high affinities for the twoother subtypes containing β2 subunits, α2β2 and α3β2 nAChRs. Incontrast, the binding affinities of these compounds for nAChR subtypescontaining β4 subunits are much lower than those for their β2 containingcounterparts. As shown in FIGS. 14 and 15, the selectivity of thesecompounds for α4β2 receptors over α3β4 receptors (K_(i) ratio) were veryhigh, ranging from 5,400 times (VMY-2-117) to 87,000 times (VMY-2-123),much greater than that of nicotine or varenicline. Similar to the lowaffinities for α3β4 receptors, these compounds showed very lowaffinities for α7 homomeric receptors.

After VMY-2-95 showed a promising binding profile, we synthesized a fewof its analogs to understand consequences of substitutions on thebenzene ring. We had substitutions at position 2, 3 or 4 to generateVMY-2-101, 105, 109, 123 and 127. These singly substituted analogs havebinding profiles similar to that of VMY-2-95 (FIG. 14). The dualsubstituted analogs, VMY-2-113, 131, 135 and 139, also showed highaffinities and selectivity for α4β2 receptors. However, the installationof CF₃ at position 3 decreased the binding affinity for the α4β2receptor, as shown by the binding profile of VMY-2-117.

Interestingly, two compounds of series 2, VMY-2-161 (azetidine replacewith cyclobutane) and VMY-2-177 (ring opened analogue of azetidine),showed very low binding affinities to nAChRs (FIG. 15), indicating thatthe azetidine ring is important for the high affinity and selectivitybinding profile. In addition, the (S)—N-methyl compound VMY-2-205 bindsnearly 25 times weaker than the corresponding (S)—N—H VMY-2-95,suggesting that N—H provides a potentially important H-bond interaction.For comparing the difference between stereotypes, VMY-2-191, which isthe (R)-form of VMY-2-95, was synthesized. The binding profile ofVMY-2-191 is similar to that of VMY-2-95, though VMY-2-191 has aslightly lower affinity for α4β2 receptors than VMY-2-95.

Binding Affinities for Targets Other than nAChRs.

To determine affinities of the lead compound, VMY-2-95, for targetsother than neuronal nAChRs, we tested VMY-2-95 in binding assays using41 other targets, including many CNS receptors and transporters. Asshown in FIG. 18, the preliminary binding assays using a singleconcentration of VMY-2-95 at 10 μM generated 32 “miss” (less than 50%inhibition of bindings by specific labeled ligands) and 9 “hit” (morethan 50% inhibition of bindings). The Ki values of VMY-2-95 at these 9targets were determined by performing secondary binding assays using aseries of concentrations of VMY-2-95. As shown in FIG. 19, the compoundhas low binding affinities for these targets. Furthermore, the bindingaffinity of VMY-2-95 for α4β2 nAChRs is at least 3,000 times higher thanthat for any of those 9 targets.

Effects of VMY-2-95 on Functions of nAChRs

The functional effect of the lead compound, VMY-2-95, was assessed bymeasuring agonist-stimulated ⁸⁶Rb⁺ efflux from stably transfected cellsexpressing nAChRs, either human α4β2 subtype or rat 03134 subtype. Itsability to desensitize nAChRs was determined by measuringnicotine-stimulated ⁸⁶Rb⁺ efflux after cells were preincubated withVMY-2-95 for 10 min. For comparison, we also examined three othernicotinic ligands in the same manner, Sazetidine-A, varenicline and(−)-nicotine.

As shown in FIG. 17, VMY-2-95 did not show any detectable agonistactivity at rat α3β4 nAChRs. At human α4β2 receptors, the compoundshowed clear agonist activity with EC₅₀ value as 8.6 nM. However, itsefficacy for activating the α4β2 receptors is very low, only 26% of themaximal stimulation by nicotine. In the parallel experiments,varenicline showed near-full agonist activity at α3β4 nAChRs but partialagonist activity at α4β2 receptors.

It is important to note that all four compounds studied, VMY-2-95,Sazetidine-A, varenicline and nicotine, inhibited nicotine activation ofα4β2 receptors after preincubated with the cells for 10 min. As shown inFIG. 17, VMY-2-95 potently desensitized 414132 receptor function with anIC_(50(10″)) value of 16 nM, which is similar to that of Sazetidine-Abut significantly lower than those of varenicline and nicotine. Incontrast, the IC_(50(10″)) value of VMY-2-95 in desensitizing α3β4nAChRs was higher than 10,000 nM, which is more than 600 times higherthan its IC_(50(10″)) value in desensitizing α4β2 nAChRs.

Effects of VMY-2-95 on Nicotine Self-Administration in Rats.

We studied acute effects of VMY-2-95 on nicotine self-administration inrats. As shown in FIG. 6, VMY-2-95 significantly (F(3,42)=3.36, p<0.05)decreased intravenous nicotine self-administration in a dose-dependentmanner. The 3 mg/kg dose of VMY-2-95 caused a significant (p<0.025)decrease in the number of nicotine infusions compared with vehicle.However, at the two lower doses studied (0.3 and 1 mg/kg), VMY-2-95 didnot produce significant effects. The effects of VMY-2-95 appeared tobecome more pronounced during the second phase of treatment (Data notshown). These data demonstrate that similar to Saz-A, VMY-2-95 at 3mg/kg effectively reduces nicotine self-administration in rats (Levin etal., 2010).

It is interesting that the locomotor activity of rats was significantlyincreased at the three doses of VMY-2-95 tested (FIG. 10). Therefore, itis highly unlikely the effect of VMY-2-95 in decreasing nicotineself-administration is resulted from a sedative effect. Furthermore, thedata indicate that the effect in locomotor activities does not correlateto the effect in nicotine self-administration, as the peak of increasedlocomotor activity was at 1 mg/kg, at which there was no significantreduction of nicotine self-administration.

Binding Affinities for Targets Other than nAChRs

To determine affinities of the lead compound, VMY-2-95, for targetsother than neuronal nAChRs, we tested VMY-2-95 in binding assays using41 other targets, including many CNS receptors and transporters. Asshown in FIG. 18, the preliminary binding assays using a singleconcentration of VMY-2-95 at 10 μM generated 32 “miss” (less than 50%inhibition of bindings by specific labeled ligands) and 9 “hit” (morethan 50% inhibition of bindings). The K_(i) values of VMY-2-95 at these9 targets were determined by performing secondary binding assays using aseries of concentrations of VMY-2-95. As shown in FIG. 19, the compoundhas low binding affinities for these targets. Furthermore, the bindingaffinity of VMY-2-95 for (14132 nAChRs is at least 3,000 times higherthan that for any of those 9 targets.

Comparison of Molecular Recognition of VMY-2-95 by Receptor inStructural Models with Those of Nicotine, Varenicline and Saz-A.

To better understand the binding interactions between α4β2 nAChR andVMY-2-95, molecular models of VMY-2-95 with the α4β2 nAChRs wereconstructed (FIG. 8), and overlaid with (−)-nicotine, varenicline, andsaz-A (FIG. 9). The docked structural models of the α4β2 nAChR/ligandcomplex reveal that the binding mode interactions are slightly differentfrom one another as reported. To provide a consistent model with thenicotine-AChBP (PDB: 1UW6), the docked positions of compounds wereremodeled using a step-by-step manual docking methodology withrestrained molecular dynamics (MD) simulations followed by energyminimization. In the restrained MD simulations, the optimum van derWaals and H-bond distance constraints was set between the ligand and theα4β2 nAChR ligand binding domain residues. The final binding complex isdepicted in FIGS. 8 and 9.

VMY-2-95, (−)-nicotine, varenicline and Saz-A are buried at the aromaticrich residues such as W147α, W55β, Y91α, Y188α, Y195α, and F117β (FIGS.8 and 9) and occupied a similar binding region. However, the relativeorientations of the compounds in the α4β2 nAChR binding site wereslightly different. This may be due to conformational adjustments insidethe binding site. The azetidine group in saz-A and VMY-2-95 formsstacking interactions with the amino acid W147α (FIGS. 8 and 9).However, the pyridine ring occupies a slightly different position tocompensate for the conformational entropy penalty due to isomericconstraints. The hydroxy group of saz-A forms a hydrogen bond with Y188αwhereas this residue forms hydrogen bond with the pyrazine ring nitrogenof varenicline, and this hydrogen bond is absent in (−)-nicotine.VMY-2-95, and saz-A have hydrophobic groups extending from the pyridinering, forming additional favorable hydrophobic interactions with K76β,K77β, Y112β, V109β, F117β, and L119β. These interactions can be comparedwith the interactions of varenicline involving V109β, F117β, and L119β(FIGS. 8 and 9). Although saz-A and VMY-2-95 have a similar pattern ofinteraction, the benzene ring of the ethynylbenzene of VMY-2-95 form astronger stacking and lipophilic interaction with K76β, K77β, Y112β, andV109β than the corresponding hex-5-yn-1-ol group of saz-A. Moreover, the3-(2-phenylethynyl) pyridine group of VMY-2-95 is more rigid and mayrequire a smaller penalty in conformational entropy as compared to theflexible 6-(3-pyridyl) hex-5-yn-1-ol group of saz-A (FIGS. 8 and 9).These additional interactions and favorable entropy penalty may likelyconfer affinity to VMY-2-95. The overlay of VMY-2-95 with saz-A,varenicline, and (−)-nicotine (FIG. 9) reveals that VMY-2-95 forms morefavorable hydrophobic interactions with the α4β2 nAChR and occupiesdifferent receptor space that may be critical for selectivity.

Physicochemistry Properties of the New Ligands

Several key physicochemical parameters of compounds may have importantroles in influencing the blood-brain barrier (BBB) penetration of CNSdrugs, including molecular weight (MW), polar surface area (PSA) andlipophilicity (clogP). These parameters of all series 1 compounds werecalculated and presented in FIG. 20.

In general CNS drugs have a MW≦450-Da. All the compounds in this reportwill have molecular weights less than 450 Da. In addition, all thesecompounds have clogP values <5, suggesting a reasonable probability ofgood oral absorption and intestinal permeability.

In general, a Polar surface area (PSA) less than 60 Å² is predictive fora compound to penetrate BBB well. As shown in FIG. 20, all the compoundsin series 1 have PSA values smaller than 60 Å².

We also calculated the log BB, which is a parameter commonly used toexpress the extent of a drug passing through the blood-brain barrier.Several QSAR model were developed to calculate the log BB. In thisstudy, we predicted the log BB values for compounds in series 1. Ingeneral a log BB value greater than zero is a favorable factor for BBBpenetration. The log BB values of all compounds in series 1 were in thepositive range.

Ligand efficiency (LE) is an important metric in drug discovery and hasbeen used to measure the relationship of biological activity (affinity)with molecular size. LE is the ratio of the free energy of binding overthe number of heavy atoms in a molecule. LE is a useful optimizationtool to evaluate a ligand's ability to effectively bind to the targetedprotein. Considering the binding affinity (IQ of the compounds in series1, we calculated the LE. All compounds in series 1 have a LE value inthe range of 0.7 kcal/mole (a LE≧0.3 is favorable, FIG. 20) and suggestthat these compounds are optimized for receptor occupancy.

Tobacco use and nicotine addiction impose a huge health and economicburden. To date, there are only three classes of medications that havebeen approved by the U.S. Food and Drug Administration for smokingcessation: nicotine replacement therapy, bupropion and varenicline.Among the three, varenicline (Chantix©) is considered superior in termsof relative efficacy. However, the percentage of subjects who remainedsmoke-free for 12 months following treatment with varenicline fell to˜22%. Moreover, although varenicline appears to be safe for most people,exacerbation of schizophrenia and manic episodes associated withtreatment with varenicline have been reported. In addition, based on arecent report, the FDA issued a “notification”, warning that vareniclinemay be associated with increased adverse cardiovascular events,including angina and heart attack. More commonly, nearly 30% ofparticipants taking varenicline in clinical trials reported nausea and18% reported vomiting.

Although varenicline was developed as a partial agonist at α4β2 nAChRs,it is also nearly a full agonist at α3β4 nAChRs, which predominate inautonomic ganglia and brainstem autonomic centers, as well as at α7nAChRs, another important subtype in brain. More recently, vareniclinewas found to be a potent agonist of the human 5-hydroxytryptamine₃receptors (5-HT₃). The side effects of varenicline are most likely to bemediated through its actions at receptors other than α4β2 nAChRs,including α3β4* nAChRs, α7 nAChRS and 5-HT₃ receptors.

Given the grave health and economic consequences of smoking there is anobvious great need for significant improvement in the existing smokingcessation therapies. In 2006, resulted from our study of Sazetidine-A(Saz-A), we proposed a new strategy to develop novel nicotinictherapeutics, including smoking cessation aids, based on their abilityto selectively desensitize α4β2 nAChRs. Saz-A is a novel nAChR ligandthat is highly selective for α4β2 receptors. In binding assays, Saz-Ahas a much higher binding affinity for α4β2 nAChRs than for α3β4 or α7receptors. After cells are exposed to it for 10 min, Saz-A potently(IC_(50(10′))=12 nM) inhibits nicotine-stimulated function mediated byα4β2 nAChRs (FIG. 17). Consistent with its high selectivity for α4β2receptor Saz-A at concentrations up to 10 μM had no effect on thefunction of rat α3β4 nAChRs (FIG. 17). This is consistent with thehypothesis that Saz-A selectively desensitizes α4β2 receptors.

The high potency of Saz-A to desensitize α4β2 nAChRs in cells in vitrosuggested that it would produce important effects similar to some ofthose produced by nicotine, but with much more receptor selectivity. Itwould therefore potentially be a drug candidate to help people overcomeaddiction to nicotine and to treat other CNS disorders. Since 2007,Saz-A has been shown to have efficacy in quite a few animal behavioralmodels.

In this study, we have developed a new class of selective α4β2 nAChRligands. These compounds maintain the excellent pharmacological propertyprofile of Saz-A with improved physicochemistry properties.

As shown in FIG. 14, in binding studies, similar to Saz-A, VMY-2-95 andanalogs are highly selective to α4β2 nAChRs over α3β4 and α7 subtypes.As assessed in ⁸⁶Rb¹ efflux assays, compared to nicotine, VMY-2-95 hasless than 30% of efficacy in activating human α4β2 nAChRs. As the sameas Saz-A, the compound does not show agonist activity at rat α3β4nAChRs. VMY-2-95 potently and selectively desensitizes α4β2 nAChRs. Itspotency for desensitization is close to that of Saz-A but much strongerthan those of varenicline and nicotine.

VMY-2-95 and other compounds in series 1 have more favorable values ofphysicochemistry parameters, such as lower PSA and higher clogP thanthose of Saz-A, indicating the possibility that they may have better BBBpenetration and higher brain distribution.

Consistent with the excellent pharmacological property profile andimproved physicochemistry properties, in the initial in vivo studies(FIGS. 5 and 6), VMY-2-95 significantly reduces nicotineself-administration at 3 mg/kg in the rat model, showing promise as asmoking cessation aid.

To elucidate SAR for VMY-2-95, in addition to compounds in series 1(VMY-2-95 through VMY-2-139), we also synthesized four compounds inseries 2, including VMY-2-161, VMY-2-177, VMY-2-191 and VMY-2-205. Thevery low binding affinities of VMY-2-161 and VMY-2-177 may be anindication that the azetidine ring is important for good bindingproperty profiles. The binding affinity of (S)—N-methyl compoundVMY-2-205 is 25 times lower than that of corresponding (S)—N-H VMY-2-95.It is conceivable that the N—H provides a potentially important H-bondinteraction. It is interesting that the two enantiomers, VMY-2-191 andVMY-2-95, showing a similar binding profile though VMY-2-191 has aslightly lower affinity for α4β2 receptors than VMY-2-95.

The binding model of α4β2 nAChR with VMY-2-95 suggests similar occupancyof the binding pocket as that of nicotine and varenicline. Moreover, thephenylethynyl group at C-5 position of pyridine in VMY-2-95 occupies apotentially critical space in the pocket and forms favorable hydrophobicinteractions with α4β2 nAChR (FIGS. 8 and 9).

In screening against 41 targets other than nAChRs (FIG. 18), VMY-2-95does not have high binding affinity for any of these targets. Thecompound either is not a “hit” in primary binding assays, or showing atleast 3,000-fold lower binding affinities than that for α4β2 nAChRs insecondary binding assays (FIG. 19,). It is important to note thatVMY-2-95 is not a “hit” at 5-HT₃ receptors, which mediate some ofvarenicline's adverse side effects, including nausea.

INCORPORATION BY REFERENCE

All of the U.S. patents and U.S. published patent applications citedherein are hereby incorporated by reference.

EQUIVALENTS

Those skilled in the art will recognize, or be able to ascertain usingno more than routine experimentation, many equivalents to the specificembodiments of the invention described herein. Such equivalents areintended to be encompassed by the following claims.

1. A compound of formula (I) or a pharmaceutically acceptable saltthereof

wherein: R represents, independently for each occurrence, halogen, C1-C6alkyl, allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, ortrifluoro-C1-C4 alkyl; R₁ represents hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl; R₂, R₃, and R₄ independently represent hydrogen, C1-C6alkyl, allyl, or C3-C6 cycloalkyl, or C1-C6 alkyl substituted with atleast one fluorine; m is an integer ranging from 1 to 3; n is an integerselected from 1 and 2; and z is an integer ranging from 0 to 5; with theproviso that the compound of formula (I) is not

2.-14. (canceled)
 15. A compound of formula (II) or a pharmaceuticallyacceptable salt thereof

wherein: R represents, independently for each occurrence, halogen, C1-C6alkyl, allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, ortrifluoro-C1-C4 alkyl; R₁ represents hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl; R₂, R₃, and R₄ independently represent hydrogen, C1-C6alkyl, allyl, or C3-C6 cycloalkyl, or C1-C6 alkyl substituted with atleast one fluorine; Y is O, S, or N(R^(C)); R^(C) is hydrogen, C1-C6alkyl, or allyloxycarbonyl; m is an integer ranging from 1 to 3; n is aninteger selected from 1 and 2; and z is an integer ranging from 0 to 5.16.-26. (canceled)
 27. A compound of formula (III) or a pharmaceuticallyacceptable salt thereof

wherein: R₁ represents hydrogen, C1-C6 alkyl, allyl, or C3-C6cycloalkyl; R₂, R₃, and R₄ independently represent hydrogen, C1-C6alkyl, allyl, or C3-C6 cycloalkyl, or C1-C6 alkyl substituted with atleast one fluorine; R₅ represents

Y represents —C≡C— or O; A represents hydrogen, halogen, methyl, C2-C6alkyl, methoxy, hydroxy, amino, trifluoromethyl, isopropyl, or t-butyl;W represents O, S, or N(R^(D)); R^(D) represents hydrogen, C1-C6 alkyl,or allyloxycarbonyl; m is an integer ranging from 1 to 3; and n is aninteger selected from 1 and
 2. 28.-35. (canceled)
 36. A compound offormula (IV) or a pharmaceutically acceptable salt thereof

wherein: R represents, independently for each occurrence, halogen, C1-C6alkyl, allyl, C1-C6 alkyloxy, amino, hydroxyl, nitro, cyano, ortrifluoro-C1-C4 alkyl; R₁ represents

R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R_(h) represents C1-C6 alkyl, C3-C6 cycloalkyl, aryl, or C1-C6 alkylsubstituted with at least one fluorine; m is an integer ranging from 1to 3; n is an integer selected from 1 and 2; and z is an integer rangingfrom 0 to
 5. 37.-44. (canceled)
 45. A compound of formula (V) or apharmaceutically acceptable salt thereof

wherein: R₁ represents

R₂, R₃, and R₄ independently represent hydrogen, C1-C6 alkyl, allyl, orC3-C6 cycloalkyl, or C1-C6 alkyl substituted with at least one fluorine;R_(h) represents C1-C6 alkyl, C3-C6 cycloalkyl, aryl, or C1-C6 alkylsubstituted with at least one fluorine; R₅ represents

Y represents —C≡C— or O; A represents hydrogen, halogen, methyl, C2-C6alkyl, methoxy, hydroxy, amino, trifluoromethyl, isopropyl, or t-butyl;W represents O, S, or N(R^(D)); R^(D) represents hydrogen, C1-C6 alkyl,or allyloxycarbonyl; m is an integer ranging from 1 to 3; and n is aninteger selected from 1 and
 2. 46-48. (canceled)
 49. A pharmaceuticalcomposition, comprising a compound of claim 1; and a pharmaceuticallyacceptable carrier.
 50. A method of treating a disorder selected fromthe group consisting of addiction, pain, obesity, schizophrenia,epilepsy, mania and manic depression, anxiety, Alzheimer's disease,learning deficit, cognition deficit, attention deficit, memory loss,Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD),Parkinson's disease, Huntington's disease, Tourette's syndrome,amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury,dyskinesias, obsessive compulsive disorder, chemical substance abuse,alcoholism, memory deficit, pseudodementia, Ganser's syndrome, migrainepain, bulimia, premenstrual syndrome or late luteal phase syndrome,tobacco abuse, post-traumatic syndrome, social phobia, chronic fatiguesyndrome, premature ejaculation, erectile difficulty, anorexia nervosa,autism, mutism, trichotillomania, hypothermia, and disorders of sleep,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of claim
 1. 51.-52. (canceled)
 53. Apharmaceutical composition, comprising a compound of claim 15; and apharmaceutically acceptable carrier.
 54. A method of treating a disorderselected from the group consisting of addiction, pain, obesity,schizophrenia, epilepsy, mania and manic depression, anxiety,Alzheimer's disease, learning deficit, cognition deficit, attentiondeficit, memory loss, Lewy Body Dementia, Attention DeficitHyperactivity Disorder (ADHD), Parkinson's disease, Huntington'sdisease, Tourette's syndrome, amyotrophic lateral sclerosis,inflammation, stroke, spinal cord injury, dyskinesias, obsessivecompulsive disorder, chemical substance abuse, alcoholism, memorydeficit, pseudodementia, Ganser's syndrome, migraine pain, bulimia,premenstrual syndrome or late luteal phase syndrome, tobacco abuse,post-traumatic syndrome, social phobia, chronic fatigue syndrome,premature ejaculation, erectile difficulty, anorexia nervosa, autism,mutism, trichotillomania, hypothermia, and disorders of sleep,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of claim
 15. 55. A pharmaceuticalcomposition, comprising a compound of claim 27; and a pharmaceuticallyacceptable carrier.
 56. A method of treating a disorder selected fromthe group consisting of addiction, pain, obesity, schizophrenia,epilepsy, mania and manic depression, anxiety, Alzheimer's disease,learning deficit, cognition deficit, attention deficit, memory loss,Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD),Parkinson's disease, Huntington's disease, Tourette's syndrome,amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury,dyskinesias, obsessive compulsive disorder, chemical substance abuse,alcoholism, memory deficit, pseudodementia, Ganser's syndrome, migrainepain, bulimia, premenstrual syndrome or late luteal phase syndrome,tobacco abuse, post-traumatic syndrome, social phobia, chronic fatiguesyndrome, premature ejaculation, erectile difficulty, anorexia nervosa,autism, mutism, trichotillomania, hypothermia, and disorders of sleep,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of claim
 27. 57. A pharmaceuticalcomposition, comprising a compound of claim 36; and a pharmaceuticallyacceptable carrier.
 58. A method of treating a disorder selected fromthe group consisting of addiction, pain, obesity, schizophrenia,epilepsy, mania and manic depression, anxiety, Alzheimer's disease,learning deficit, cognition deficit, attention deficit, memory loss,Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD),Parkinson's disease, Huntington's disease, Tourette's syndrome,amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury,dyskinesias, obsessive compulsive disorder, chemical substance abuse,alcoholism, memory deficit, pseudodementia, Ganser's syndrome, migrainepain, bulimia, premenstrual syndrome or late luteal phase syndrome,tobacco abuse, post-traumatic syndrome, social phobia, chronic fatiguesyndrome, premature ejaculation, erectile difficulty, anorexia nervosa,autism, mutism, trichotillomania, hypothermia, and disorders of sleep,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of claim
 36. 59. A pharmaceuticalcomposition, comprising a compound of claim 45; and a pharmaceuticallyacceptable carrier.
 60. A method of treating a disorder selected fromthe group consisting of addiction, pain, obesity, schizophrenia,epilepsy, mania and manic depression, anxiety, Alzheimer's disease,learning deficit, cognition deficit, attention deficit, memory loss,Lewy Body Dementia, Attention Deficit Hyperactivity Disorder (ADHD),Parkinson's disease, Huntington's disease, Tourette's syndrome,amyotrophic lateral sclerosis, inflammation, stroke, spinal cord injury,dyskinesias, obsessive compulsive disorder, chemical substance abuse,alcoholism, memory deficit, pseudodementia, Ganser's syndrome, migrainepain, bulimia, premenstrual syndrome or late luteal phase syndrome,tobacco abuse, post-traumatic syndrome, social phobia, chronic fatiguesyndrome, premature ejaculation, erectile difficulty, anorexia nervosa,autism, mutism, trichotillomania, hypothermia, and disorders of sleep,comprising administering to a subject in need thereof a therapeuticallyeffective amount of a compound of claim 45.